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Sample records for shiga toxin production

  1. Oxidative Stress in Shiga Toxin Production by Enterohemorrhagic Escherichia coli

    Directory of Open Access Journals (Sweden)

    Katarzyna Licznerska

    2016-01-01

    Full Text Available Virulence of enterohemorrhagic Escherichia coli (EHEC strains depends on production of Shiga toxins. These toxins are encoded in genomes of lambdoid bacteriophages (Shiga toxin-converting phages, present in EHEC cells as prophages. The genes coding for Shiga toxins are silent in lysogenic bacteria, and prophage induction is necessary for their efficient expression and toxin production. Under laboratory conditions, treatment with UV light or antibiotics interfering with DNA replication are commonly used to induce lambdoid prophages. Since such conditions are unlikely to occur in human intestine, various research groups searched for other factors or agents that might induce Shiga toxin-converting prophages. Among other conditions, it was reported that treatment with H2O2 caused induction of these prophages, though with efficiency significantly lower relative to UV-irradiation or mitomycin C treatment. A molecular mechanism of this phenomenon has been proposed. It appears that the oxidative stress represents natural conditions provoking induction of Shiga toxin-converting prophages as a consequence of H2O2 excretion by either neutrophils in infected humans or protist predators outside human body. Finally, the recently proposed biological role of Shiga toxin production is described in this paper, and the “bacterial altruism” and “Trojan Horse” hypotheses, which are connected to the oxidative stress, are discussed.

  2. Entry of Shiga toxin into cells

    DEFF Research Database (Denmark)

    Sandvig, Kirsten; van Deurs, Bo

    1994-01-01

    Cellebiologi, Shiga toxin, receptors, glycolipids, endocytosis, trans-Golgi network, endoplasmic reticulum, retrograde transport......Cellebiologi, Shiga toxin, receptors, glycolipids, endocytosis, trans-Golgi network, endoplasmic reticulum, retrograde transport...

  3. Altruism of Shiga toxin-producing Escherichia coli: recent hypothesis versus experimental results

    Directory of Open Access Journals (Sweden)

    Joanna M Los

    2013-01-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC may cause bloody diarrhea and hemorrhagic colitis, with subsequent systemic disease. Since genes coding for Shiga toxins (stx genes are located on lambdoid prophages, their effective production occurs only after prophage induction. Such induction and subsequent lytic development of Shiga toxin-converting bacteriophages results not only in production of toxic proteins, but also in the lysis (and thus, the death of the host cell. Therefore, one may ask the question: what is the benefit for bacteria to produce the toxin if they die due to phage production and subsequent cell lysis? Recently, a hypothesis was proposed (simultaneously but independently by two research groups that STEC may benefit from Shiga toxin production as a result of toxin-dependent killing of eukaryotic cells such as unicellular predators or human leukocytes. This hypothesis could make sense only if we assume that prophage induction (and production of the toxin occurs only in a small fraction of bacterial cells, thus, a few members of the population are sacrificed for the benefit of the rest, providing an example of ‘bacterial altruism’. However, various reports indicating that the frequency of spontaneous induction of Shiga toxin-converting prophages is higher than that of other lambdoid prophages might seem to contradict the for-mentioned model. On the other hand, analysis of recently published results, discussed here, indicated that the efficiency of prophage excision under conditions that may likely occur in the natural habitat of STEC is sufficiently low to ensure survival of a large fraction of the bacterial host. A molecular mechanism by which partial prophage induction may occur is proposed. We conclude that the published data supports the proposed model of bacterial ‘altruism’ where prophage induction occurs at a low enough frequency to render toxin production a positive selective force on the general STEC population.

  4. Isolation of Shiga toxin-producing Escherichia coli harboring variant Shiga toxin genes from seafood

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    Sreepriya Prakasan

    2018-03-01

    Full Text Available Background and Aim: Shiga toxin-producing Escherichia coli (STEC are important pathogens of global significance. STEC are responsible for numerous food-borne outbreaks worldwide and their presence in food is a potential health hazard. The objective of the present study was to determine the incidence of STEC in fresh seafood in Mumbai, India, and to characterize STEC with respect to their virulence determinants. Materials and Methods: A total of 368 E. coli were isolated from 39 fresh seafood samples (18 finfish and 21 shellfish using culture-based methods. The isolates were screened by polymerase chain reaction (PCR for the genes commonly associated with STEC. The variant Shiga toxin genes were confirmed by Southern blotting and hybridization followed by DNA sequencing. Results: One or more Shiga toxins genes were detected in 61 isolates. Of 39 samples analyzed, 10 (25.64% samples harbored STEC. Other virulence genes, namely, eaeA (coding for an intimin and hlyA (hemolysin A were detected in 43 and 15 seafood isolates, respectively. The variant stx1 genes from 6 isolates were sequenced, five of which were found to be stx1d variants, while one sequence varied considerably from known stx1 sequences. Southern hybridization and DNA sequence analysis suggested putative Shiga toxin variant genes (stx2 in at least 3 other isolates. Conclusion: The results of this study showed the occurrence of STEC in seafood harboring one or more Shiga toxin genes. The detection of STEC by PCR may be hampered due to the presence of variant genes such as the stx1d in STEC. This is the first report of stx1d gene in STEC isolated from Indian seafood.

  5. Emergence of Escherichia coli encoding Shiga toxin 2f in human Shiga toxin-producing E-coli (STEC) infections in the Netherlands, January 2008 to December 2011

    NARCIS (Netherlands)

    Friesema, I.; van der Zwaluw, K.; Schuurman, T.; Kooistra-Smid, M.; Franz, E.; van Duynhoven, Y.; van Pelt, W.

    2014-01-01

    The Shiga toxins of Shiga toxin-producing Escherichia coli (STEC) can be divided into Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) with several sub-variants. Variant Stx(2f) is one of the latest described, but has been rarely associated with symptomatic human infections. In the enhanced STEC

  6. 77 FR 9888 - Shiga Toxin-Producing Escherichia coli

    Science.gov (United States)

    2012-02-21

    ... Toxin-Producing Escherichia coli in Certain Raw Beef Products AGENCY: Food Safety and Inspection Service... toxin-producing Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, and O145). This new date..., that are contaminated with Shiga toxin-producing Escherichia coli (STEC) O26, O45, O103, O111, O121...

  7. Shiga Toxin (Stx) Gene Detection and Verotoxigenic Potentials of ...

    African Journals Online (AJOL)

    DR-AMADI

    Nigerian Journal of Basic and Applied Science (June, 2016), 24(1): 98-105 .... dangerous pathogenic shiga- toxin producing E. coli from the food product. Consequent .... Table 3: Vero Toxin Analysis of non – 0157 E. coli Isolates From Nono Sold in Nigeria. City .... receptors in their plasma membranes and will detect all ...

  8. Shiga toxin induces membrane reorganization and formation of long range lipid order

    DEFF Research Database (Denmark)

    Solovyeva, Vita; Johannes, Ludger; Simonsen, Adam Cohen

    2015-01-01

    membrane reordering. When Shiga toxin was added above the lipid chain melting temperature, the toxin interaction with the membrane induced rearrangement and clustering of Gb3 lipids that resulted in the long range order and alignment of lipids in gel domains. The toxin induced redistribution of Gb3 lipids...... inside gel domains is governed by the temperature at which Shiga toxin was added to the membrane: above or below the phase transition. The temperature is thus one of the critical factors controlling lipid organization and texture in the presence of Shiga toxin. Lipid chain ordering imposed by Shiga toxin...... binding can be another factor driving the reconstruction of lipid organization and crystallization of lipids inside gel domains....

  9. Detection of Shiga Toxins by Lateral Flow Assay

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    Kathryn H. Ching

    2015-04-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC produce shiga toxins (Stxs that can cause human disease and death. The contamination of food products with STEC represents a food safety problem that necessitates rapid and effective detection strategies to mitigate risk. In this manuscript, we report the development of a colorimetric lateral flow assay (LFA for the rapid detection of Stxs in <10 min using a pair of monoclonal antibodies that bind epitopes common to Stx1 and six Stx2 variants. This LFA provides a rapid and sensitive test for the detection of Stxs directly from STEC culture supernatants or at risk food samples with a 0.1 ng/mL limit of detection (LOD for Stx2a. This Stx LFA is applicable for use in the rapid evaluation of Stx production from cultured E. coli strains or as a tool to augment current methods as part of food safety testing.

  10. Escherichia coli Shiga Toxin Mechanisms of Action in Renal Disease

    Directory of Open Access Journals (Sweden)

    Tom G. Obrig

    2010-12-01

    Full Text Available Shiga toxin-producing Escherichia coli is a contaminant of food and water that in humans causes a diarrheal prodrome followed by more severe disease of the kidneys and an array of symptoms of the central nervous system. The systemic disease is a complex referred to as diarrhea-associated hemolytic uremic syndrome (D+HUS. D+HUS is characterized by thrombocytopenia, microangiopathic hemolytic anemia, and acute renal failure. This review focuses on the renal aspects of D+HUS. Current knowledge of this renal disease is derived from a combination of human samples, animal models of D+HUS, and interaction of Shiga toxin with isolated renal cell types. Shiga toxin is a multi-subunit protein complex that binds to a glycosphingolipid receptor, Gb3, on select eukaryotic cell types. Location of Gb3 in the kidney is predictive of the sites of action of Shiga toxin. However, the toxin is cytotoxic to some, but not all cell types that express Gb3. It also can cause apoptosis or generate an inflammatory response in some cells. Together, this myriad of results is responsible for D+HUS disease.

  11. Shigella Sonnei and Shiga Toxin

    Centers for Disease Control (CDC) Podcasts

    2016-07-28

    Katherine Lamba, an infectious disease epidemiologist with the California Department of Public Health, discusses Shiga Toxin producing Shigella sonnei.  Created: 7/28/2016 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 7/28/2016.

  12. Detection of Shiga toxins genes by Multiplex PCR in clinical samples

    Directory of Open Access Journals (Sweden)

    2013-09-01

    Full Text Available Background: Different methods have been used for detection of shiga toxins; such as,  cell culture, ELISA, and RFPLA. However, all of these methods suffer from high cost, time-consumption and relatively low sensitivity. In this study we used Multiplex PCR method for detection of genes encoding shiga toxins. Material and Methods: In this study, 63 clinical samples were obtained from positive cultures of Shigella and E. coli O157, from Bahman 1391 until Ordibehesht 1392 in Mazandaran province. Initial confirmation of shiga toxins producing bacteria was performed by biochemical and serological methods. After DNA extraction, detection of stx1 and stx2 genes was accomplished by multiplex PCR.  For confirmation of the PCR amplicon, DNA sequencing was used. Antibiotic sensitivity tests were performed by disk diffusion method. Results:  Among the positive strains, 13 strains contained stx2 genes, 4 strains contained Stx/Stx1 genes and 4 strains harbored both Stx/Stx1 and Stx2. The DNA extracted from other Gram-negative bacteria was not protected by the relevant parts of these toxins. Sequencing of the amplified fragments indicated the correct toxin sequences.  The sensitivity for identification of Stx/Stx1 gene was 1.56 pg/ µl and for Stx2 was 1.08 pg/µl. The toxin positive strains were all sensitive to Cefixime, Gentamicin, Amikacin, Ceftriaxone, and Nitrofurantoin. Conclusion: This method is fast and accurate for detection of bacteria producing shiga toxin and can be used to identify different types of shiga toxin.

  13. A High-Throughput, Precipitating Colorimetric Sandwich ELISA Microarray for Shiga Toxins

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    Andrew Gehring

    2014-06-01

    Full Text Available Shiga toxins 1 and 2 (Stx1 and Stx2 from Shiga toxin-producing E. coli (STEC bacteria were simultaneously detected with a newly developed, high-throughput antibody microarray platform. The proteinaceous toxins were immobilized and sandwiched between biorecognition elements (monoclonal antibodies and pooled horseradish peroxidase (HRP-conjugated monoclonal antibodies. Following the reaction of HRP with the precipitating chromogenic substrate (metal enhanced 3,3-diaminobenzidine tetrahydrochloride or DAB, the formation of a colored product was quantitatively measured with an inexpensive flatbed page scanner. The colorimetric ELISA microarray was demonstrated to detect Stx1 and Stx2 at levels as low as ~4.5 ng/mL within ~2 h of total assay time with a narrow linear dynamic range of ~1–2 orders of magnitude and saturation levels well above background. Stx1 and/or Stx2 produced by various strains of STEC were also detected following the treatment of cultured cells with mitomycin C (a toxin-inducing antibiotic and/or B-PER (a cell-disrupting, protein extraction reagent. Semi-quantitative detection of Shiga toxins was demonstrated to be sporadic among various STEC strains following incubation with mitomycin C; however, further reaction with B-PER generally resulted in the detection of or increased detection of Stx1, relative to Stx2, produced by STECs inoculated into either axenic broth culture or culture broth containing ground beef.

  14. Comparison of non-O157 Shiga toxin-producing E. coli detection systems

    Science.gov (United States)

    Category: methodology improvements Objective: To identify strengths and weaknesses of commercial Shiga toxin-producing E. coli detection systems and kits in a side by side fashion. Experimental Design: Three commercial Shiga toxin-producing E. coli detection tests (BAX, GDS, and GeneDisc) and two t...

  15. NNDSS - Table II. Shiga toxin to Shigellosis

    Data.gov (United States)

    U.S. Department of Health & Human Services — NNDSS - Table II. Shiga toxin to Shigellosis - 2015. In this Table, provisional cases of selected notifiable diseases (≥1,000 cases reported during the preceding...

  16. NNDSS - Table II. Shiga toxin to Shigellosis

    Data.gov (United States)

    U.S. Department of Health & Human Services — NNDSS - Table II. Shiga toxin to Shigellosis - 2016. In this Table, provisional* cases of selected† notifiable diseases (≥1,000 cases reported during the preceding...

  17. Literature Reference for Shiga and Shiga-like Toxin (Journal of Clinical Microbiology. 2007. 45(10): 3377–3380)

    Science.gov (United States)

    This procedure describes a rapid optical immunoassay for the detection of Stx-1 and Stx-2 using a commercially available kit for presumptive analysis of aerosol, solid, particulate, liquid and water samples contaminated with Shiga and Shiga-like toxin.

  18. Characterizing RecA-independent induction of Shiga toxin2-encoding phages by EDTA treatment.

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    Lejla Imamovic

    Full Text Available BACKGROUND: The bacteriophage life cycle has an important role in Shiga toxin (Stx expression. The induction of Shiga toxin-encoding phages (Stx phages increases toxin production as a result of replication of the phage genome, and phage lysis of the host cell also provides a means of Stx toxin to exit the cell. Previous studies suggested that prophage induction might also occur in the absence of SOS response, independently of RecA. METHODOLOGY/PRINCIPAL FINDINGS: The influence of EDTA on RecA-independent Stx2 phage induction was assessed, in laboratory lysogens and in EHEC strains carrying Stx2 phages in their genome, by Real-Time PCR. RecA-independent mechanisms described for phage λ induction (RcsA and DsrA were not involved in Stx2 phage induction. In addition, mutations in the pathway for the stress response of the bacterial envelope to EDTA did not contribute to Stx2 phage induction. The effect of EDTA on Stx phage induction is due to its chelating properties, which was also confirmed by the use of citrate, another chelating agent. Our results indicate that EDTA affects Stx2 phage induction by disruption of the bacterial outer membrane due to chelation of Mg(2+. In all the conditions evaluated, the pH value had a decisive role in Stx2 phage induction. CONCLUSIONS/SIGNIFICANCE: Chelating agents, such as EDTA and citrate, induce Stx phages, which raises concerns due to their frequent use in food and pharmaceutical products. This study contributes to our understanding of the phenomenon of induction and release of Stx phages as an important factor in the pathogenicity of Shiga toxin-producing Escherichia coli (STEC and in the emergence of new pathogenic strains.

  19. Phylogenetic diversity and similarity of active sites of Shiga toxin (stx) in Shiga toxin-producing Escherichia coli (STEC) isolates from humans and animals.

    Science.gov (United States)

    Asakura, H; Makino, S; Kobori, H; Watarai, M; Shirahata, T; Ikeda, T; Takeshi, K

    2001-08-01

    Nucleotide sequences of Shiga toxin (Stx) genes in STEC from various origins were determined and characterized by phylogenetic analysis based on Shiga toxin (Stx) with those deposited in GenBank. The phylogenetic trees placed Stx1 and Stx2 into two and five groups respectively, and indicated that Stx1 in sheep-origin STEC were placed into a different group from those in other STEC, and that Stx2 of deer-origin STEC also belonged to the unique group and appeared to be distantly related to human-origin STEC. On the other hand, Stx of STEC isolated from cattle, seagulls and flies were closely related to those of human-origin STEC. Such a diversity of Stx suggested that STEC might be widely disseminated in many animal species, and be dependent on their host species or their habitat. In addition, the active sites in both toxins were compared; the active sites in both subunits of Stx in all the animal-origin STEC were identical to those in human-origin STEC, suggesting that all the toxin of STEC from animals might be also cytotoxic, and therefore, such animal-origin STEC might have potential pathogenicity for humans.

  20. Evaluation of ELISA tests specific for Shiga toxin 1 and 2 in food and water samples

    Science.gov (United States)

    Two enzyme-linked immunosorbent assay (ELISA) kits were evaluated for their effectiveness in detecting and differentiating between Shiga toxin 1 and 2 (Stx1 and Stx2) produced by Shiga toxin-producing E. coli (STEC) inoculated into food and water samples. Each kit incorporated monoclonal antibodies ...

  1. Prevalence and antimicrobial sensitivity of Shiga-toxin-producing ...

    African Journals Online (AJOL)

    Background: Escherichia coli is among the most common causes of diarrhoea in children below five years of age in developing countries. Diarrhoeal diseases rank the second most common cause of morbidity and mortality in developing countries. Here we report the magnitude of Shiga toxin-producing Escherichia coli ...

  2. Detection and characterization of Shiga toxin-producing Escherichia coli in game meat and ready-to-eat meat products.

    Science.gov (United States)

    Díaz-Sánchez, S; Sánchez, S; Sánchez, M; Herrera-León, S; Hanning, I; Vidal, D

    2012-11-15

    A total of 142 samples of game meat and ready-to-eat meat products from red deer and wild boar were analysed in order to assess the presence of Shiga toxin-producing Escherichia coli (STEC). Shiga-toxin encoding genes (stx genes) were detected by PCR in 36 (25.4%) of the samples and STEC was isolated from 8 (5.6%) of the same samples. None of the samples tested positive for E. coli O157:H7. Four different serotypes were found among the 8 STEC isolates, with serotype O27:H30 being predominant (62.5%, 5/8). The PCR assay indicated the presence of the stx2 gene in all of the STEC isolates and further subtyping resulted in detection of three different subtypes: stx2a, stx2b and stx2g. The only stx1-positive isolate was further subtyped as stx1c. The ehxA gene was detected in 3 (37.5%) of the isolates and none of them contained the eae gene. All STEC isolates were sensitive to the 13 antibiotics tested. Some isolates possessed serotypes and virulence gene profiles previously associated with STEC infections in humans. The isolation of a STEC strain carrying the stx2a subtype from a ready-to-eat meat product from deer suggests the role of these products as a potential source of STEC infections in humans. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. Mechanism of Shiga Toxin Clustering on Membranes

    DEFF Research Database (Denmark)

    Pezeshkian, Weria; Gao, Haifei; Arumugam, Senthil

    2017-01-01

    between them. The precise mechanism by which this clustering occurs remains poorly defined. Here, we used vesicle and cell systems and computer simulations to show that line tension due to curvature, height, or compositional mismatch, and lipid or solvent depletion cannot drive the clustering of Shiga...... toxin molecules. By contrast, in coarse-grained computer simulations, a correlation was found between clustering and toxin nanoparticle-driven suppression of membrane fluctuations, and experimentally we observed that clustering required the toxin molecules to be tightly bound to the membrane surface...... molecules (several nanometers), and persist even beyond. This force is predicted to operate between manufactured nanoparticles providing they are sufficiently rigid and tightly bound to the plasma membrane, thereby suggesting a route for the targeting of nanoparticles to cells for biomedical applications....

  4. Translational errors in expression of Shiga toxin from pathogenic Escherichia coli as measured by MALDI-TOF-TOF and Orbitrap mass spectrometry

    Science.gov (United States)

    Introduction: Shiga toxin (Stx) is an AB5 toxin expressed by Shiga toxin-producing E. coli (STEC) and Shigella dysenteriae. The Stx holotoxin attaches to surface receptors of eukaryotic cells. After cellular envelopment, the toxin disrupts ribosomal protein synthesis causing cell death. Variations i...

  5. Failure of manganese to protect from Shiga toxin.

    Directory of Open Access Journals (Sweden)

    Marsha A Gaston

    Full Text Available Shiga toxin (Stx, the main virulence factor of Shiga toxin producing Escherichia coli, is a major public health threat, causing hemorrhagic colitis and hemolytic uremic syndrome. Currently, there are no approved therapeutics for these infections; however manganese has been reported to provide protection from the Stx1 variant isolated from Shigella dysenteriae (Stx1-S both in vitro and in vivo. We investigated the efficacy of manganese protection from Stx1-S and the more potent Stx2a isoform, using experimental systems well-established for studying Stx: in vitro responses of Vero monkey kidney cells, and in vivo toxicity to CD-1 outbred mice. Manganese treatment at the reported therapeutic concentration was toxic to Vero cells in culture and to CD-1 mice. At lower manganese concentrations that were better tolerated, we observed no protection from Stx1-S or Stx2a toxicity. The ability of manganese to prevent the effects of Stx may be particular to certain cell lines, mouse strains, or may only be manifested at high, potentially toxic manganese concentrations.

  6. Possible mistranslation of Shiga toxin from pathogenic Escherichia coli as measured by MALDI-TOF and Orbitrap mass spectrometry

    Science.gov (United States)

    RATIONALE: Shiga toxin-producing Escherichia coli (STEC) are often subjected to DNA damaging antibiotics during culturing in order to elicit the bacterial SOS response and up-regulation of bacteriophage-encoded proteins including Shiga toxin (Stx). However, such antibiotic exposure and stress may al...

  7. Glycan Encapsulated Gold Nanoparticles Selectively Inhibit Shiga Toxins 1 and 2

    OpenAIRE

    Kulkarni, Ashish A.; Fuller-Schaefer, Cynthia; Korman, Henry; Weiss, Alison A.; Iyer, Suri S.

    2010-01-01

    Shiga toxins (Stx) released by Escherichia coli O157:H7 and Shigella dysentriae, cause life-threatening conditions that include hemolytic-uremic syndrome (HUS), kidney failure and neurological complications. Cellular entry is mediated by the B subunit of the AB5 toxin, which recognizes cell surface glycolipids present in lipid raft like structures. We developed gold glyconanoparticles that present a multivalent display similar to the cell surface glycolipids to compete for these toxins. These...

  8. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    OpenAIRE

    Parma, Y. R.; Chacana, P. A.; Lucchesi, P. M. A.; Rogé, A.; Granobles Velandia, C. V.; Krüger, A.; Parma, A. E.; Fernández-Miyakawa, M. E.

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC), a subset of Shiga toxin producing E. coli (STEC) is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic-uremic syndrome (HUS). Regardless of serotype, Shiga toxins (Stx1 and/or Stx2) are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx) was developed using anti-St...

  9. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    OpenAIRE

    Yanil R Parma; Pablo A Chacana; Paula María Alejandra Lucchesi; Ariel eRoge; Claudia V Granobles Velandia; Alejandra eKrüger; Alejandra eKrüger; Alberto E. Parma; Mariano Enrique Fernandez Miyakawa

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC), a subset of Shiga toxin producing E. coli (STEC) is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic uremic syndrome (HUS). Regardless of serotype, Shiga toxins (Stx1 and/or Stx2) are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx) was developed using anti-St...

  10. Sub-Lethal Dose of Shiga toxin 2 from Enterohemorrhagic Escherichia coli Affects Balance and Cerebellar Cythoarquitecture.

    Directory of Open Access Journals (Sweden)

    Luciana eD’Alessio

    2016-02-01

    Full Text Available Shiga toxin producing Escherichia coli may damage the central nervous system before or concomitantly to manifested hemolytic uremic syndrome symptoms. The cerebellum is frequently damaged during this syndrome, however the deleterious effects of Shiga toxin 2 has never been integrally reported by ultrastructural, physiological and behavioral means. The aim of this study was to determine the cerebellar compromise after intravenous administration of a sub-lethal dose of Shiga toxin 2 by measuring the cerebellar blood brain barrier permeability, behavioral task of cerebellar functionality (inclined plane test, and ultrastructural analysis (transmission electron microscope. Intravenous administration of vehicle (control group, sub-lethal dose of 0.5 ηg and 1 ηg of Stx2 per mouse were tested for behavioral and ultrastructural studies. A set of three independent experiments were performed for each study (n=6. Blood–Brain Barrier resulted damaged and consequently its permeability was significantly increased. Lower scores obtained in the inclined plane task denoted poor cerebellar functionality in comparison to their controls. The most significant lower score was obtained after 5 days of 1ηg of toxin administration. Transmission electron microscope micrographs from the Stx2-treated groups showed neurons with a progressive neurodegenerative condition in a dose dependent manner. As sub-lethal intravenous Shiga toxin 2 altered the blood brain barrier permeability in the cerebellum the toxin penetrated the cerebellar parenchyma and produced cell damaged with significant functional implications in the test balance.

  11. Shiga Toxin (Stx) Gene Detection and Verotoxigenic Potentials of ...

    African Journals Online (AJOL)

    Non-0157 Escherichia coli, isolated from Nono (fermented fresh cow milk) sampled from four major Nigerian cities, namely, Abuja, Benin City, Lagos and Onitsha were investigated for the presence shiga toxins (stx1 and stx2) genes using PCR technique and for their verotoxigenic potentials using tissue culture assay on ...

  12. Food Forensics: Using Mass Spectrometry To Detect Foodborne Protein Contaminants, as Exemplified by Shiga Toxin Variants and Prion Strains.

    Science.gov (United States)

    Silva, Christopher J

    2018-06-13

    Food forensicists need a variety of tools to detect the many possible food contaminants. As a result of its analytical flexibility, mass spectrometry is one of those tools. Use of the multiple reaction monitoring (MRM) method expands its use to quantitation as well as detection of infectious proteins (prions) and protein toxins, such as Shiga toxins. The sample processing steps inactivate prions and Shiga toxins; the proteins are digested with proteases to yield peptides suitable for MRM-based analysis. Prions are detected by their distinct physicochemical properties and differential covalent modification. Shiga toxin analysis is based on detecting peptides derived from the five identical binding B subunits comprising the toxin. 15 N-labeled internal standards are prepared from cloned proteins. These examples illustrate the power of MRM, in that the same instrument can be used to safely detect and quantitate protein toxins, prions, and small molecules that might contaminate our food.

  13. Expression and purification of recombinant Shiga toxin 2B from ...

    African Journals Online (AJOL)

    Expression and purification of recombinant Shiga toxin 2B from Escherichia coli O157:H7. ... (SDS-PAGE) and StxB2 yield was 450 μg ml-1 confirmed by Bradford assay. Recombinant Stx2B protein was produced in highly pure yield using ...

  14. Shiga toxin-producing Escherichia coli in slaughtered pigs and pork products

    Directory of Open Access Journals (Sweden)

    Lia Bardasi

    2017-05-01

    Full Text Available During the years 2015-2016, 83 faecal samples were collected at slaughter from pigs reared in farms located in Central- Northern Italy. During the years 2014-2016 a total of 562 pork products [465 not-ready-to-eat (NRTE and 97 ready-to-eat (RTE products] were collected from retail outlets, large retailers and processing plants. The samples were analysed according to ISO TS 13136:2012. Out of 83 swine faecal samples, 77 (92.8% resulted stx-positive by real time polymerase chain reaction (PCR, 5 stx2+ and 1 stx1+ Shiga toxin-producing Escherichia coli (STEC strains were isolated. Among the 465 NRTE samples, 65 (14.0% resulted stx-positive by real time PCR and 7 stx2+ STEC strains were isolated. The stx2 gene was detected more frequently than the stx1 gene both in faecal samples (90.4 vs 8.4% and in NRTE pork products (13.3 vs 1.3%. All the RTE samples included in the analysis resulted stxnegative. Among the samples resulted positive for stx and eae genes, serogroup-associated genes were detected at high frequency: O26 resulted the most frequent in faecal samples (81.3% and O145 in pork products (88.1%. The O157 serogroup resulted positive in 83.3 and 78.1% of pork products and faecal samples, respectively. Despite the frequent detection by real time PCR of genes indicating the possible presence of STEC strains belonging to the six serogroups, the bacteriological step did not confirm the isolation of any such strains.

  15. Shiga Toxin in Enterohemorrhagic E.coli: regulation and novel antivirulence strategies

    Directory of Open Access Journals (Sweden)

    Vanessa eSperandio

    2012-06-01

    Full Text Available Enterohemorrhagic Escherichia coli O157:H7 infects about 73,000 people annually in the USA and is a major cause of outbreaks of bloody diarrhea worldwide, and. In 5 to 7% of the cases, the person infected develops the potentially fatal sequelae hemolytic uremic syndrome (HUS, characterized by acute kidney failure. A hallmark of EHEC pathogenesis and cause of HUS is the production of Shiga toxin (Stx. Stx was first described by Kiyoshi Shiga in Shigella dysenterae serotype I and later discovered in EHEC, and it has been linked to HUS since 1983. Many factors regulate the production of Stx, including temperature, growth phase, antibiotics, reactive oxygen species and quorum sensing. Currently, there is no effective treatment or prophylaxis for HUS. Since the use of antibiotics is not advised to treat EHEC infections because it triggers Stx production, alternative antibacterial strategies need to be developed. Quorum sensing inhibitors represent a novel class of antibacterial compounds, which have the advantage of not interfering on bacterial growth, thereby without selective pressure that can lead to appearance of resistant strains. In this review, we discuss factors that regulate Stx production in EHEC as well as novel strategies to fight Stx and minimize development to HUS in EHEC-infected patients.

  16. Human mannose-binding lectin inhibitor prevents Shiga toxin-induced renal injury

    DEFF Research Database (Denmark)

    Ozaki, Masayuki; Kang, Yulin; Tan, Ying Siow

    2016-01-01

    Hemolytic uremic syndrome caused by Shiga toxin-producing Escherichia coli (STEC HUS) is a worldwide endemic problem, and its pathophysiology is not fully elucidated. Here we tested whether the mannose-binding lectin (MBL2), an initiating factor of lectin complement pathway activation, plays a cr...

  17. Inhibition of Shiga toxin 2 (Stx2) in apple juices and its resistance to pasteurization.

    Science.gov (United States)

    Rasooly, Reuven; Do, Paula M; Levin, Carol E; Friedman, Mendel

    2010-06-01

    In the present study, we evaluated Shiga toxin (Stx2) activity in apple juices by measuring a decrease in dehydrogenase activity of Vero cells with the microculture tetrazolium (MTT) assay. Freshly prepared juice from Red Delicious apples and Golden Delicious apples inhibited the biological activity of the bacterial toxin Stx2 produced by E. coli O157:H7 strains. Studies with immunomagnetic beads bearing specific antibodies against the toxin revealed that Stx2 activity was restored when removed from the apple juice. SDS gel electrophoresis revealed no difference (P pasteurize apple juice, but lost all activity when exposed to 100 degrees C for 5 min. The results suggest that pasteurization of apple juice used to inactivate E. coli O157:H7 has no effect on Stx2, and that food-compatible and safe antitoxin compounds can be used to inhibit the biological activity of the Shiga toxin.

  18. Isolation, genotyping, and antimicrobial resistance of zoonotic shiga toxin-producing escherichia coli

    Science.gov (United States)

    Shiga toxin-producing Escherichia coli (STEC) is an enteric pathogen linked to outbreaks of human gastroenteritis with diverse clinical spectra. Traditional culture and isolation methods, including selective enrichment and differential plating, have enabled the effective recovery of STEC. Ruminants ...

  19. Lipid reorganization induced by Shiga toxin clustering on planar membranes.

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    Barbara Windschiegl

    Full Text Available The homopentameric B-subunit of bacterial protein Shiga toxin (STxB binds to the glycolipid Gb(3 in plasma membranes, which is the initial step for entering cells by a clathrin-independent mechanism. It has been suggested that protein clustering and lipid reorganization determine toxin uptake into cells. Here, we elucidated the molecular requirements for STxB induced Gb(3 clustering and for the proposed lipid reorganization in planar membranes. The influence of binding site III of the B-subunit as well as the Gb(3 lipid structure was investigated by means of high resolution methods such as fluorescence and scanning force microscopy. STxB was found to form protein clusters on homogenous 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC/cholesterol/Gb(3 (65:30:5 bilayers. In contrast, membranes composed of DOPC/cholesterol/sphingomyelin/Gb(3 (40:35:20:5 phase separate into a liquid ordered and liquid disordered phase. Dependent on the fatty acid composition of Gb(3, STxB-Gb(3 complexes organize within the liquid ordered phase upon protein binding. Our findings suggest that STxB is capable of forming a new membrane phase that is characterized by lipid compaction. The significance of this finding is discussed in the context of Shiga toxin-induced formation of endocytic membrane invaginations.

  20. Antimicrobial Drug-Resistant Shiga Toxin-Producing Escherichia coli Infections, Michigan, USA.

    Science.gov (United States)

    Mukherjee, Sanjana; Mosci, Rebekah E; Anderson, Chase M; Snyder, Brian A; Collins, James; Rudrik, James T; Manning, Shannon D

    2017-09-01

    High frequencies of antimicrobial drug resistance were observed in O157 and non-O157 Shiga toxin-producing E. coli strains recovered from patients in Michigan during 2010-2014. Resistance was more common in non-O157 strains and independently associated with hospitalization, indicating that resistance could contribute to more severe disease outcomes.

  1. Phylogeny and disease association of Shiga toxin-producing Escherichia coli O91

    NARCIS (Netherlands)

    Mellmann, Alexander; Fruth, Angelika; Friedrich, Alexander W; Wieler, Lothar H; Harmsen, Dag; Werber, Dirk; Middendorf, Barbara; Bielaszewska, Martina; Karch, Helge

    The diversity and relatedness of 100 Shiga toxin-producing Escherichia coli O91 isolates from different patients were examined by multilocus sequence typing. We identified 10 specific sequence types (ST) and 4 distinct clonal groups. ST442 was significantly associated with hemolytic uremic syndrome.

  2. Thermal inactivation of Escherichia coli 0157:H7 (ECOH) and non-0157 Shiga toxin-producing E.coli (STEC)in mechanically tenderized veal

    Science.gov (United States)

    We quantified thermal destruction of Shiga toxin-producing Escherichia coli O157:H7 (ECOH) and Shiga toxin-producing non-O157 E. coli (STEC) cells within mechanically tenderized veal cutlets following cooking on an electric skillet. For each of five trials, flattened veal cutlets (ca. 71.6 g; ca. 1/...

  3. New monoclonal antibodies against a novel subtype of Shiga toxin 1 produced by Enterobacter cloacae and their use in analysis of human serum

    Science.gov (United States)

    Shiga toxin (Stx) is a major virulence factor for several bacterial pathogens that cause potentially fatal illness, including Escherichia coli and Shigella spp. The continual emergence of new subtypes of Stxs presents challenges in clinical diagnosis of infections caused by Shiga toxin-producing org...

  4. Cationic antimicrobial peptides inactivate Shiga toxin-encoding bacteriophages

    Science.gov (United States)

    Del Cogliano, Manuel E.; Hollmann, Axel; Martinez, Melina; Semorile, Liliana; Ghiringhelli, Pablo D.; Maffía, Paulo C.; Bentancor, Leticia V.

    2017-12-01

    Shiga toxin (Stx) is the principal virulence factor during Shiga toxin-producing Escherichia coli (STEC) infections. We have previously reported the inactivation of bacteriophage encoding Stx after treatment with chitosan, a linear polysaccharide polymer with cationic properties. Cationic antimicrobial peptides (cAMPs) are short linear aminoacidic sequences, with a positive net charge, which display bactericidal or bacteriostatic activity against a wide range of bacterial species. They are promising novel antibiotics since they have shown bactericidal effects against multiresistant bacteria. To evaluate whether cationic properties are responsible for bacteriophage inactivation, we tested seven cationic peptides with proven antimicrobial activity as anti-bacteriophage agents, and one random sequence cationic peptide with no antimicrobial activity as a control. We observed bacteriophage inactivation after incubation with five cAMPs, but no inactivating activity was observed with the random sequence cationic peptide or with the non alpha helical cAMP Omiganan. Finally, to confirm peptide-bacteriophage interaction, zeta potential was analyzed by following changes on bacteriophage surface charges after peptide incubation. According to our results we could propose that: 1) direct interaction of peptides with phage is a necessary step for bacteriophage inactivation, 2) cationic properties are necessary but not sufficient for bacteriophage inactivation, and 3) inactivation by cationic peptides could be sequence (or structure) specific. Overall our data suggest that these peptides could be considered a new family of molecules potentially useful to decrease bacteriophage replication and Stx expression.

  5. Cationic Antimicrobial Peptides Inactivate Shiga Toxin-Encoding Bacteriophages

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    Manuel E. Del Cogliano

    2017-12-01

    Full Text Available Shiga toxin (Stx is the principal virulence factor during Shiga toxin-producing Escherichia coli (STEC infections. We have previously reported the inactivation of bacteriophage encoding Stx after treatment with chitosan, a linear polysaccharide polymer with cationic properties. Cationic antimicrobial peptides (cAMPs are short linear aminoacidic sequences, with a positive net charge, which display bactericidal or bacteriostatic activity against a wide range of bacterial species. They are promising novel antibiotics since they have shown bactericidal effects against multiresistant bacteria. To evaluate whether cationic properties are responsible for bacteriophage inactivation, we tested seven cationic peptides with proven antimicrobial activity as anti-bacteriophage agents, and one random sequence cationic peptide with no antimicrobial activity as a control. We observed bacteriophage inactivation after incubation with five cAMPs, but no inactivating activity was observed with the random sequence cationic peptide or with the non-alpha helical cAMP Omiganan. Finally, to confirm peptide-bacteriophage interaction, zeta potential was analyzed by following changes on bacteriophage surface charges after peptide incubation. According to our results we could propose that: (1 direct interaction of peptides with phage is a necessary step for bacteriophage inactivation, (2 cationic properties are necessary but not sufficient for bacteriophage inactivation, and (3 inactivation by cationic peptides could be sequence (or structure specific. Overall our data suggest that these peptides could be considered a new family of molecules potentially useful to decrease bacteriophage replication and Stx expression.

  6. Shiga toxin-producing Escherichia coli in humans and the food chain in Bangladesh

    NARCIS (Netherlands)

    Islam, M.A.

    2009-01-01

    Shiga toxin-producing Escherichia coli (STEC) are significant pathogenic bacteria that can cause severe gastrointestinal diseases and also the hemolytic-uremic syndrome. Domestic ruminants appear to be the main reservoirs of these organisms. Although Bangladesh is an endemic zone for diarrhea caused

  7. Differences in Shiga toxin and phage production among stx2g-positive STEC strains

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    Claudia Viviana Granobles Velandia

    2012-06-01

    Full Text Available Shigatoxigenic E. coli (STEC are characterized by the production of Shiga toxins (Stx encoded by temperate bacteriophages. Stx production is linked to the induction of the phage lytic cycle. Several stx variants have been described and differentially associated with the risk of developing severe illness.The variant named stx2g was first identified in a STEC strain isolated from the faeces of healthy cattle. Analysis of stx2g-positive strains isolated from humans, animals and environmental sources have shown that they have a close relationship. In this study, stx2g-positive STEC isolated from cattle were analyzed for phage and Stx production, with the aim to relate the results to differences observed in cytotoxicity.The presence of inducible phages was assessed by analyzing the bacterial growth/lysis curves and also by plaque assay. Bacterial growth curves in the absence of induction were similar for all isolates, however, notably differed among induced cultures. The two strains that clearly evidenced bacteriolysis under this condition also showed higher phage titers in plaque assays. However, only the phage plaques produced by one of these strains (FB 62 hybridized with a stx2-probe. Furthermore, the production of Stx was evaluated by EIA and Western immunoblotting in overnight supernatants. By EIA, we detected Stx only in supernatants of FB 62, with a higher signal with induced than in uninduced cultures. By immunoblotting, Stx2 could be detected after induction in all stx2g-positive isolates, but with lower amounts of Stx2B subunit in those supernatants where phages could not be detected.Taking into account all the results, several differences could be found among stx2g-positive strains. The strain with the highest cytotoxic titer showed higher levels of stx2-phages and toxin production by EIA, and the opposite was observed for strains that previously showed low cytotoxic titers, confirming that in stx2g-positive strains Stx production is

  8. The epithelial cell cytoskeleton and intracellular trafficking. I. Shiga toxin B-subunit system: retrograde transport, intracellular vectorization, and more.

    Science.gov (United States)

    Johannes, Ludger

    2002-07-01

    Many intracellular transport routes are still little explored. This is particularly true for retrograde transport between the plasma membrane and the endoplasmic reticulum. Shiga toxin B subunit has become a powerful tool to study this pathway, and recent advances on the molecular mechanisms of transport in the retrograde route and on its physiological function(s) are summarized. Furthermore, it is discussed how the study of retrograde transport of Shiga toxin B subunit allows one to design new methods for the intracellular delivery of therapeutic compounds.

  9. A plant-based oral vaccine to protect against systemic intoxication by Shiga toxin type 2

    OpenAIRE

    Wen, Sharon X.; Teel, Louise D.; Judge, Nicole A.; O’Brien, Alison D.

    2006-01-01

    Hemolytic uremic syndrome, the leading cause of kidney failure in children, often follows infection with enterohemorrhagic Escherichia coli and is mediated by the Shiga type toxins, particularly type 2 (Stx2), produced by such strains. The challenge in protecting against this life-threatening syndrome is to stimulate an immune response at the site of infection while also protecting against Shiga intoxication at distal sites such as the kidney. As one approach to meeting this challenge, we sou...

  10. PCR Based Detection of Shiga Toxin Producing E. coli in Commercial Poultry and Related Environments

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    Homaira Anzum Himi

    2015-02-01

    Full Text Available Shiga toxin (Stx-producing E. coli (STEC is the most important foodborne pathogen which is the causal agent of mild diarrhea, bloody diarrhea, hemolytic-uremic syndrome (HUS in human. The present study was designed to determine the prevalence and identification of Shiga toxin (Stx-producing E. coli in poultry, detection of its source of infection in poultry and transmission pattern to human. For this purpose a total of 150 samples (cloacal swab-60, feed -15, water-15 and egg -60 were collected and analyzed in bacteriology laboratory by cultured in different bacteriological media followed by gram’s staining, biochemical tests and Polymerase Chain reaction (PCR. The PCR was performed by targeting 16s rRNA gene and shiga toxin producing gene in E. coli. Out of 150 collected samples, E. coli was found in 81 (54% samples. Presence of E. coli was 100% in both feed (n=15 and egg (n=60, whereas 10% in cloacal swab (n=6. Water samples were totally free of E. coli. The stx2 gene was detected in all samples whether all samples were negative for stx1 gene. The study revealed that, poultry feed acts as a source of E. coli infection in poultry, which may be transmitted to environment and human via meat or eggs. Antibiotic sensitivity test revealed that isolated bacteria were highly sensitive to Ciprofloxacin.

  11. Detection, Characterization and Typing of Shiga toxin-producing Escherichia coli.

    OpenAIRE

    Brendon David Parsons; Nathan eZelyas; Byron M Berenger; Linda eChui; Linda eChui

    2016-01-01

    Shiga toxin-producing Escherichia coli (STEC) are responsible for gastrointestinal diseases reported in numerous outbreaks around the world. Given the public health importance of STEC, effective detection, characterization and typing is critical to any medical laboratory system. While non-O157 serotypes account for the majority of STEC infections, frontline microbiology laboratories may only screen for STEC using O157-specific agar-based methods. As a result, non-O157 STEC infections are sign...

  12. Effect of antibiotics on cellular stress generated in Shiga toxin-producing Escherichia coli O157:H7 and non-O157 biofilms.

    Science.gov (United States)

    Angel Villegas, Natalia; Baronetti, José; Albesa, Inés; Etcheverría, Analía; Becerra, M Cecilia; Padola, Nora L; Paraje, M Gabriela

    2015-10-01

    Shiga toxin-producing Escherichia coli (STEC) are important food-borne pathogens, with the main virulence factor of this bacterium being its capacity to secrete Shiga toxins (Stxs). Therefore, the use of certain antibiotics for the treatment of this infection, which induces the liberation of Stxs, is controversial. Reactive oxygen and nitrogen species are also involved in the pathogenesis of different diseases. The purpose of this study was to analyze the effects of antibiotics on biofilms of STEC and the relationships between cellular stress and the release of Stx. To this end, biofilms of reference and clinical strains were treated with antibiotics (ciprofloxacin, fosfomycin and rifaximin) and the production of oxidants, the antioxidant defense system and toxin release were evaluated. Ciprofloxacin altered the prooxidant-antioxidant balance, with a decrease of oxidant metabolites and an increase of superoxide dismutase and catalase activity, being associated with high-levels of Stx production. Furthermore, inhibition of oxidative stress by exogenous antioxidants was correlated with a reduction in the liberation of Stx, indicating the participation of this phenomenon in the release of this toxin. In contrast, fosfomycin and rifaximin produced less alteration with a minimal production of Stx. Our data show that treatment of biofilm-STEC with these antibiotics induces oxidative stress-mediated release of Stx. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Identification of human-pathogenic strains of Shiga toxin-producing Escherichia coli from food by a combination of serotyping and molecular typing of Shiga toxin genes.

    Science.gov (United States)

    Beutin, Lothar; Miko, Angelika; Krause, Gladys; Pries, Karin; Haby, Sabine; Steege, Katja; Albrecht, Nadine

    2007-08-01

    We examined 219 Shiga toxin-producing Escherichia coli (STEC) strains from meat, milk, and cheese samples collected in Germany between 2005 and 2006. All strains were investigated for their serotypes and for genetic variants of Shiga toxins 1 and 2 (Stx1 and Stx2). stx(1) or variant genes were detected in 88 (40.2%) strains and stx(2) and variants in 177 (80.8%) strains. Typing of stx genes was performed by stx-specific PCRs and by analysis of restriction fragment length polymorphisms (RFLP) of PCR products. Major genotypes of the Stx1 (stx(1), stx(1c), and stx(1d)) and the Stx2 (stx(2), stx(2d), stx(2-O118), stx(2e), and stx(2g)) families were detected, and multiple types of stx genes coexisted frequently in STEC strains. Only 1.8% of the STEC strains from food belonged to the classical enterohemorrhagic E. coli (EHEC) types O26:H11, O103:H2, and O157:H7, and only 5.0% of the STEC strains from food were positive for the eae gene, which is a virulence trait of classical EHEC. In contrast, 95 (43.4%) of the food-borne STEC strains carried stx(2) and/or mucus-activatable stx(2d) genes, an indicator for potential high virulence of STEC for humans. Most of these strains belonged to serotypes associated with severe illness in humans, such as O22:H8, O91:H21, O113:H21, O174:H2, and O174:H21. stx(2) and stx(2d) STEC strains were found frequently in milk and beef products. Other stx types were associated more frequently with pork (stx(2e)), lamb, and wildlife meat (stx(1c)). The combination of serotyping and stx genotyping was found useful for identification and for assignment of food-borne STEC to groups with potential lower and higher levels of virulence for humans.

  14. Comparative genomics and stx phage characterization of LEE-negative Shiga toxin-producing Escherichia coli

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    Susan Renee Steyert

    2012-11-01

    Full Text Available Infection by Escherichia coli and Shigella species are among the leading causes of death due to diarrheal disease in the world. Shiga toxin producing Escherichia coli (STEC that do not encode the locus of enterocyte effacement (LEE-negative STEC often possess Shiga toxin gene variants and have been isolated from humans and a variety of animal sources. In this study, we compare the genomes of nine LEE-negative STEC harboring various stx alleles with four complete reference LEE-positive STEC isolates. Compared to a representative collection of prototype E. coli and Shigella isolates representing each of the pathotypes, the whole genome phylogeny demonstrated that these isolates are diverse. Whole genome comparative analysis of the 13 genomes revealed that in addition to the absence of the LEE pathogenicity island, phage encoded genes including non-LEE encoded effectors, were absent from all nine LEE-negative STEC genomes. Several plasmid-encoded virulence factors reportedly identified in LEE-negative STEC isolates were identified in only a subset of the nine LEE-negative isolates further confirming the diversity of this group. In combination with whole genome analysis, we characterized the lambdoid phages harboring the various stx alleles and determined their genomic insertion sites. Although the integrase gene sequence corresponded with genomic location, it was not correlated with stx variant, further highlighting the mosaic nature of these phages. The transcription of these phages in different genomic backgrounds was examined. Expression of the Shiga toxin genes, stx1 and/or stx2, as well as the Q genes, were examined with quantitative reverse transcriptase polymerase chain reaction (qRT-PCR assays. A wide range of basal and induced toxin induction was observed. Overall, this is a first significant foray into the genome space of this unexplored group of emerging and divergent pathogens.

  15. Heterogeneity in Induction Level, Infection Ability, and Morphology of Shiga Toxin-Encoding Phages (Stx Phages) from Dairy and Human Shiga Toxin-Producing Escherichia coli O26:H11 Isolates

    Science.gov (United States)

    Bonanno, Ludivine; Petit, Marie-Agnès; Loukiadis, Estelle; Michel, Valérie

    2016-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) bacteria are foodborne pathogens responsible for diarrhea and hemolytic-uremic syndrome (HUS). Shiga toxin, the main STEC virulence factor, is encoded by the stx gene located in the genome of a bacteriophage inserted into the bacterial chromosome. The O26:H11 serotype is considered to be the second-most-significant HUS-causing serotype worldwide after O157:H7. STEC O26:H11 bacteria and their stx-negative counterparts have been detected in dairy products. They may convert from the one form to the other by loss or acquisition of Stx phages, potentially confounding food microbiological diagnostic methods based on stx gene detection. Here we investigated the diversity and mobility of Stx phages from human and dairy STEC O26:H11 strains. Evaluation of their rate of in vitro induction, occurring either spontaneously or in the presence of mitomycin C, showed that the Stx2 phages were more inducible overall than Stx1 phages. However, no correlation was found between the Stx phage levels produced and the origin of the strains tested or the phage insertion sites. Morphological analysis by electron microscopy showed that Stx phages from STEC O26:H11 displayed various shapes that were unrelated to Stx1 or Stx2 types. Finally, the levels of sensitivity of stx-negative E. coli O26:H11 to six Stx phages differed among the 17 strains tested and our attempts to convert them into STEC were unsuccessful, indicating that their lysogenization was a rare event. PMID:26826235

  16. Classification of shiga toxin-producing escherichia coli (STEC) serotypes with hyperspectral microscope imagery

    Science.gov (United States)

    Non-O157:H7 Shiga toxin-producing Escherichia coli (STEC) strains such as O26, O45, O103, O111, O121 and O145 are recognized as serious outbreak to cause human illness due to their toxicity. Since a conventional microbiological method for cell counting is laborious and time-consuming process, optica...

  17. Shiga toxin-producing Escherichia coli (STEC: principal virulence factors and epidemiology Escherichia coli produtora de toxina shiga (STEC: principais fatores de virulência e dados epidemiológicos

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    Halha Ostrensky Saridakis

    2007-10-01

    Full Text Available Shiga toxin producing Escherichia coli is an important food borne pathogen, mainly beef products, and is associated to mild and severe bloody diarrhea. In some individuals, STEC infection can progress to hemolytic-uremic syndrome (HUS, a sequela characterized by renal failure, and thrombotic thrombocytopenic purpura (TTP, with possible central nervous system involvement. Cattle, usually healthy, is the principal reservoir of STEC, although these strains have also been isolated from other domestic animals: sheep, goats, dogs, cats and pigs. The principal virulence feature, the production of Shiga toxins, is not enough to cause diseases, and other factors are considered important, as enterohemolysin and fimbrial and afimbrial adhesions production. Although human diseases associated to STEC have not been frequently reported in Brazil, their presence is frequent in cattle, as well as the correlation between serotypes found in these animals and human patients. Escherichia coli produtora de toxina Shiga (STEC é um importante patógeno veiculado por alimentos, principalmente produtos derivados de carne bovina e está associado a quadros de diarréias leves a severas e sanguinolentas. Em alguns indivíduos, a infecção por STEC pode progredir para a síndrome hemolítico-urêmica (HUS, seqüela caracterizada pela falência renal e a púrpura trombocitopênica trombótica (TTP, com possível envolvimento do sistema nervoso central. O gado bovino, geralmente saudável, é o principal reservatório de STEC, embora estas cepas também tenham sido isoladas de outros animais domésticos: ovelhas, cabras, cães, gatos e suínos. A principal característica de virulência, a produção de toxinas Shiga, não é suficiente para causar doenças e outros fatores são considerados relevantes, como a produção de enterohemolisina e de adesinas fimbriais e afimbriais. Embora as doenças humanas associadas a STEC sejam pouco descritas no Brasil, podemos observar

  18. UV-Sensitivity of Shiga Toxin-Converting Bacteriophage Virions Φ24B, 933W, P22, P27 and P32

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    Sylwia Bloch

    2015-09-01

    Full Text Available Shiga toxin-converting bacteriophages (Stx phages are present as prophages in Shiga toxin-producing Escherichia coli (STEC strains. Theses phages can be transmitted to previously non-pathogenic E. coli cells making them potential producers of Shiga toxins, as they bear genes for these toxins in their genomes. Therefore, sensitivity of Stx phage virions to various conditions is important in both natural processes of spreading of these viruses and potential prophylactic control of appearance of novel pathogenic E. coli strains. In this report we provide evidence that virions of Stx phages are significantly more sensitive to UV irradiation than bacteriophage λ. Following UV irradiation of Stx virions at the dose of 50 J/m2, their infectivity dropped by 1–3 log10, depending on the kind of phage. Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions. Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development. We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells.

  19. Inhibition of Shiga Toxin 2 (Stx2) in Apple Juices and its Resistance to Pasteurization

    Science.gov (United States)

    In the present study, we evaluated Shiga toxin (Stx2) activity in apple juices by measuring a decrease in dehydrogenase activity of Vero cells with the microculture tetrazolium (MTT) assay. Freshly prepared juice from Red Delicious apples and Golden Delicious apples inhibited the biological activity...

  20. Detection of Escherichia coli Shiga toxin-producing in viscera of animals bovine and chicken intended for human consumption

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    Zotta, Claudio Marcelo

    2016-05-01

    Full Text Available Escherichia coli producing-Shiga toxin (STEC is associated with foodborne illness (ETA. It can cause bloody diarrhea, hemorrhagic colitis, hemolytic uremic syndrome and thrombotic thrombocytopenic purpura. The aim of the study was to detect the presence of STEC in samples of organs (offal of bovine animals and chicken intended for human consumption. Between 2008-2009, 76 samples bovine entrails and 22 chicken viscera samples, were processed and underwent, as screening technique, the polymerase chain reaction (PCR for detection of multiple genes coding for the factors virulence: Shiga toxin (stx1, stx2 and rfbO157 gene coding for capsular O157 lipopolysaccharide LPS. Samples from bovine offal development showed 84.2% for coliform bacteria. These isolates showed no virulence factor that characterized as STEC or Escherichia coli O157. The chicken offal samples showed 95.5% of development for coliform bacteria, being negative for the presence of genes encoding the Shiga toxins 1 and 2 (stx1, stx2 and rfbO157 gene. While this work does not STEC was detected, the presence of coliform bacteria in the samples studied makes these foods should be considered as potentially hazardous to consume undercooked with the consequent possibility of filing ETA.

  1. Salt at concentrations relevant to meat processing enhances Shiga toxin 2 production in Escherichia coli O157:H7.

    Science.gov (United States)

    Harris, Shaun M; Yue, Wan-Fu; Olsen, Sarena A; Hu, Jia; Means, Warrie J; McCormick, Richard J; Du, Min; Zhu, Mei-Jun

    2012-10-15

    Escherichia coli (E. coli) O157:H7 remains a major food safety concern associated with meat, especially beef products. Shiga toxins (Stx) are key virulence factors produced by E. coli O157:H7 that are responsible for hemorrhagic colitis and Hemolytic Uremic Syndrome. Stx are heat stable and can be absorbed after oral ingestion. Despite the extensive study of E. coli O157:H7 survival during meat processing, little attention is paid to the production of Stx during meat processing. The objective of this study was to elucidate the effect of salt, an essential additive to processed meat, at concentrations relevant to meat processing (0%, 1%, 2%, 3%, W/V) on Stx2 production and Stx2 prophage induction by E. coli O157:H7 strains. For both E. coli O157:H7 86-24 and EDL933 strains, including 2% salt in LB broth decreased (Pmeat processing enhances Stx production, a process linked to bacterial stress response and lambdoid prophage induction. Published by Elsevier B.V.

  2. Comparison of Escherichia coli Isolates from humans, food, and farm and companion animals for presence of Shiga toxin-producing E. coli virulence markers.

    Science.gov (United States)

    Murinda, Shelton E; Nguyen, Lien T; Landers, Tippi L; Draughon, F Ann; Mathew, Alan G; Hogan, Joseph S; Smith, K Larry; Hancock, Dale D; Oliver, Stephen P

    2004-01-01

    The objective of this study was to characterize Escherichia coli isolates from dairy cows/feedlots, calves, mastitis, pigs, dogs, parrot, iguana, human disease, and food products for prevalence of Shiga toxin-producing E. coli (STEC) virulence markers. The rationale of the study was that, isolates of the same serotypes that were obtained from different sources and possessed the same marker profiles, could be cross-species transmissible. Multiplex polymerase chain reaction (PCR) was used to detect presence of genes encoding Shiga toxin 1 and 2 (stx1 and stx2), H7 flagella (flicC), enterohemolysin (hly) and intimin (eaeA) in E. coli isolates (n = 400). Shiga toxin-producing isolates were tested for production of Shiga toxins (Stx1 and Stx2 and enterohemolysin. Of the E. coli O157:H7/H- strains, 150 of 164 (mostly human, cattle, and food) isolates were stx+. Sixty-five percent of O157 STEC produced both Stx1 and Stx2; 32% and 0.7% produced Stx2 or Stx1, respectively. Ninety-eight percent of O157 STEC had sequences for genes encoding intimin and enterohemolysin. Five of 20 E. coli O111, 4 of 14 O128 and 4 of 10 O26 were stx+ . Five of 6 stx+ O26 and O111 produced Stx1, however, stx+ O128 were Stx-negative. Acid resistance (93.3%) and tellurite resistance (87.3%) were common attributes of O157 STEC, whereas, non-O157 stx+ strains exhibited 38.5% and 30.8% of the respective resistances. stx-positive isolates were mostly associated with humans and cattle, whereas, all isolates from mastitis (n = 105), and pigs, dogs, parrot and iguanas (n = 48) were stx-negative. Multiplex PCR was an effective tool for characterizing STEC pathogenic profiles and distinguished STEC O157:H7 from other STEC. Isolates from cattle and human disease shared similar toxigenic profiles, whereas isolates from other disease sources had few characteristics in common with the former isolates. These data suggest interspecies transmissibility of certain serotypes, in particular, STEC O157:H7, between

  3. Isolation and characteristics of Shiga toxin 2f-producing Escherichia coli among pigeons in Kyushu, Japan.

    Directory of Open Access Journals (Sweden)

    Koichi Murakami

    Full Text Available An increasing number of Shiga toxin 2f-producing Escherichia coli (STEC2f infections in humans are being reported in Europe, and pigeons have been suggested as a reservoir for the pathogen. In Japan, there is very little information regarding carriage of STEC2f by pigeons, prompting the need for further investigation. We collected 549 samples of pigeon droppings from 14 locations in Kyushu, Japan, to isolate STEC2f and to investigate characteristics of the isolates. Shiga toxin stx 2f gene fragments were detected by PCR in 16 (2.9% of the 549 dropping samples across four of the 14 locations. We obtained 23 STEC2f-isolates from seven of the original samples and from three pigeon dropping samples collected in an additional sampling experiment (from a total of seven locations across both sampling periods. Genotypic and phenotypic characteristics were then examined for selected isolates from each of 10 samples with pulsed-field gel electrophoresis profiles. Eight of the stx 2f gene fragments sequenced in this study were homologous to others that were identified in Europe. Some isolates also contained virulence-related genes, including lpfA O26, irp 2, and fyuA, and all of the 10 selected isolates maintained the eae, astA, and cdt genes. Moreover, five of the 10 selected isolates contained sfpA, a gene that is restricted to Shiga toxin-producing E. coli O165:H2 and sorbitol-fermenting Shiga toxin-producing E. coli O157:NM. We document serotypes O152:HNM, O128:HNM, and O145:H34 as STEC2f, which agrees with previous studies on pigeons and humans. Interestingly, O119:H21 was newly described as STEC2f. O145:H34, with sequence type 722, was described in a German study in humans and was also isolated in the current study. These results revealed that Japanese zoonotic STEC2f strains harboring several virulence-related factors may be of the same clonal complexes as some European strains. These findings provide useful information for public health

  4. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    Science.gov (United States)

    Parma, Y. R.; Chacana, P. A.; Lucchesi, P. M. A.; Rogé, A.; Granobles Velandia, C. V.; Krüger, A.; Parma, A. E.; Fernández-Miyakawa, M. E.

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC), a subset of Shiga toxin producing E. coli (STEC) is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic-uremic syndrome (HUS). Regardless of serotype, Shiga toxins (Stx1 and/or Stx2) are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx) was developed using anti-Stx2B subunit antibodies and its performance was compared with that of the Vero cell assay and a commercial immunoassay kit. Chicken IgY was used as capture antibody and a HRP-conjugated rabbit IgG as the detection antibody. The anti-Stx2B IgY was harvested from eggs laid by hens immunized with a recombinant protein fragment. Several parameters were tested in order to optimize the sandwich ELISA assay, including concentration of antibodies, type and concentration of blocking agent, and incubation temperatures. Supernatants from 42 STEC strains of different serotypes and stx variants, including stx2EDL933, stx2vha, stx2vhb, stx2g, stx1EDL933, and stx1d were tested. All Stx variants were detected by the sandwich ELISA, with a detection limit of 115 ng/ml Stx2. Twenty three strains negative for stx genes, including different bacteria species, showed no activity in Vero cell assay and produced negative results in ELISA, except for two strains. Our results show that anti-Stx2B IgY sandwich ELISA could be used in routine diagnosis as a rapid, specific and economic method for detection of Shiga toxin-producing E. coli. PMID:22919675

  5. Prevalence and characteristics of Shiga toxin-producing Escherichia coli in finishing pigs: implications on public health

    Science.gov (United States)

    Shiga toxin-producing Escherichia coli (STEC) are important food-borne pathogens, which can cause serious illnesses, including hemorrhagic colitis and hemolytic uremic syndrome. To examine if pigs are potential animal reservoirs for human STEC infections, we conducted a longitudinal cohort study in ...

  6. Evaluation of beef trim sampling methods for detection of Shiga toxin-producing Escherichia coli (STEC)

    Science.gov (United States)

    Presence of Shiga toxin-producing Escherichia coli (STEC) is a major concern in ground beef. Several methods for sampling beef trim prior to grinding are currently used in the beef industry. The purpose of this study was to determine the efficacy of the sampling methods for detecting STEC in beef ...

  7. Shiga toxin-producing Escherichia coli in Central Greece: prevalence and virulence genes of O157:H7 and non-O157 in animal feces, vegetables, and humans.

    Science.gov (United States)

    Pinaka, O; Pournaras, S; Mouchtouri, V; Plakokefalos, E; Katsiaflaka, A; Kolokythopoulou, F; Barboutsi, E; Bitsolas, N; Hadjichristodoulou, C

    2013-11-01

    In Greece, Shiga toxin-producing Escherichia coli (STEC) have only been sporadically reported. The objective of this study was to estimate the prevalence of STEC and Escherichia coli O157:H7 in farm animals, vegetables, and humans in Greece. A total number of 1,010 fecal samples were collected from farm animals (sheep, goats, cattle, chickens, pigs), 667 diarrheal samples from humans, and 60 from vegetables, which were cultured in specific media for STEC isolates. Enzyme-linked immunosorbent assay (ELISA) was used to detect toxin-producing colonies, which, subsequently, were subjected to a multiplex polymerase chain reaction (PCR) for stx1, stx2, eae, rfbE O157, and fliC h7 genes. Eighty isolates (7.9 %) from animal samples were found to produce Shiga toxin by ELISA, while by PCR, O157 STEC isolates were detected from 8 (0.8 %) samples and non-O157 STEC isolates from 43 (4.2 %) samples. STEC isolates were recovered mainly from sheep and goats, rarely from cattle, and not from pigs and chickens, suggesting that small ruminants constitute a potential risk for human infections. However, only three human specimens (0.4 %) were positive for the detection of Shiga toxins and all were PCR-negative. Similarly, all 60 vegetable samples were negative for toxin production and for toxin genes, but three samples (two roman rockets and one spinach) were positive by PCR for rfbE O157 and fliC h7 genes. These findings indicate that sheep, goats, cattle, and leafy vegetables can be a reservoir of STEC and Escherichia coli O157:H7 isolates in Greece, which are still rarely detected among humans.

  8. Inactivation of shiga toxin-producing Escherichia coli in lean ground beef by gamma irradiation

    Science.gov (United States)

    Non-O157 serovars of Shiga Toxin-producing Escherichia coli (STEC) are now responsible for over 60% of STEC induced illnesses. The majority of illnesses caused by non-O157:H7 STEC have been due to serogroups O26, O121, O103, O45, O111, and O145, “the big/top six”, which are now considered adulterant...

  9. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    Directory of Open Access Journals (Sweden)

    Yanil R Parma

    2012-06-01

    Full Text Available Enterohemorrhagic Escherichia coli (EHEC, a subset of Shiga toxin producing E. coli (STEC is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic uremic syndrome (HUS. Regardless of serotype, Shiga toxins (Stx1 and/or Stx2 are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx was developed using anti-Stx2 B subunit antibodies and its performance was compared with that of the Vero cell assay and a commercial immunoassay kit. Chicken IgY was used as capture antibody and a HRP-conjugated rabbit IgG as the detection antibody. The anti-Stx2B IgY was harvested from eggs laid by hens immunized with a recombinant protein fragment. Several parameters were tested in order to optimize the sandwich ELISA assay, including concentration of antibodies, type and concentration of blocking agent, and incubation temperatures. Supernatants from 42 STEC strains of different serotypes and stx variants, including stx2EDL933, stx2vha, stx2vhb, stx2g, stx1EDL933 and stx1d were tested. All Stx variants were detected by the sandwich ELISA, with a detection limit of 400 ng /ml Stx2. Twenty three strains negative for stx genes, including different bacteria species, showed no activity in Vero cell assay and produced negative results in ELISA, except for 2 strains. Our results show that anti-Stx2B IgY sandwich ELISA could be used in routine diagnosis as a rapid, specific and economic method for detection of Shiga toxin-producing E. coli.

  10. Shiga Toxin Increases Formation of Clathrin-Coated Pits through Syk Kinase

    DEFF Research Database (Denmark)

    Utskarpen, Audrun; Massol, Ramiro; van Deurs, Bo

    2010-01-01

    Clathrin-dependent endocytosis is a main entry mechanism for the glycolipid-binding Shiga toxin (Stx), although clathrin-independent pathways are also involved. Binding of Stx to its receptor Gb3 not only is essential for Stx retrograde transport to the endoplasmic reticulum and toxicity but also...... activates signaling through the tyrosine kinase Syk. We previously described that Syk activity is important for Stx entry, but it remained unclear how this kinase modulates endocytosis of Stx. Here we characterized the effects of Stx and Syk on clathrin-coated pit formation. We found that acute treatment...

  11. Molecular characterization and phylogeny of Shiga toxin-producing E. coli (STEC) from imported beef meat in Malaysia.

    Science.gov (United States)

    Abuelhassan, Nawal Nouridaim; Mutalib, Sahilah Abdul; Gimba, Fufa Ido; Yusoff, Wan Mohtar

    2016-09-01

    This study aimed at determining the presence and characterization of Escherichia coli and Shiga toxin-producing E. coli (STEC) from imported frozen beef meats. Seventy-four (74) frozen imported beef meat samples from two countries, India (42 samples) and Australia (32 samples), were collected and tested for E. coli. These samples were purchased from the frozen meat sections of five different supermarkets in different locations in Selangor, Malaysia, from April 2012 to October 2014. A total of 222 E. coli strains were isolated from the meat samples; 126 strains were isolated from country A (India), and 96 E. coli strains were from country of origin B (Australia), respectively. A total of 70 E. coli strains were identified and characterized. All E. coli strains were isolated into Fluorocult medium and identified using API 20E kit. All selected E. coli strains were characterized for Shiga toxin genes (stx1 and stx2). All biochemically identified E. coli in this study were further subjected to molecular detection through polymerase chain reaction (PCR) amplification and characterization using 16S ribosomal RNA (rRNA) gene of Shiga toxin-producing E. coli. Of the 70 E. coli strains, 11 strains were positive for both Shiga toxin genes (stx1 and stx2) and 11 (11/70) strains were positive for stx1 gene, while 25 (25/70) strains were positive for stx2 gene. The analysis of 16S rRNA gene of all the E. coli isolates in this study was successfully sequenced and analyzed, and based on sequence data obtained, a phylogenetic tree of the 16S rRNA gene was performed using Clustal W programme in MEGA 6.06 software. Phylogenetic tree showed that the E. coli isolates in our study cluster with the strain of E. coli isolated in other countries, which further confirm that the isolates of E. coli in this study are similar to those obtained in other studies. As a result, all the strains obtained in this study proved to be a strain of pathogenic E. coli, which may cause a serious outbreak

  12. Outbreak of Non-O157 Shiga Toxin-Producing Escherichia coli Infection from Consumption of Beef Sausage

    DEFF Research Database (Denmark)

    Ethelberg, S.; Smith, B.; Torpdahl, M.

    2009-01-01

    We describe an outbreak of Shiga toxin-producing Escherichia coli O26: H11 infection in 20 patients (median age, 2 years). The source of the infection was an organic fermented beef sausage. The source was discovered by using credit card information to obtain and compare customer transaction records...

  13. Quiescent complement in nonhuman primates during E coli Shiga toxin-induced hemolytic uremic syndrome and thrombotic microangiopathy.

    Science.gov (United States)

    Lee, Benjamin C; Mayer, Chad L; Leibowitz, Caitlin S; Stearns-Kurosawa, D J; Kurosawa, Shinichiro

    2013-08-01

    Enterohemorrhagic Escherichia coli (EHEC) produce ribosome-inactivating Shiga toxins (Stx1, Stx2) responsible for development of hemolytic uremic syndrome (HUS) and acute kidney injury (AKI). Some patients show complement activation during EHEC infection, raising the possibility of therapeutic targeting of complement for relief. Our juvenile nonhuman primate (Papio baboons) models of endotoxin-free Stx challenge exhibit full spectrum HUS, including thrombocytopenia, hemolytic anemia, and AKI with glomerular thrombotic microangiopathy. There were no significant increases in soluble terminal complement complex (C5b-9) levels after challenge with lethal Stx1 (n = 6) or Stx2 (n = 5) in plasma samples from T0 to euthanasia at 49.5 to 128 hours post-challenge. d-dimer and cell injury markers (HMGB1, histones) confirmed coagulopathy and cell injury. Thus, complement activation is not required for the development of thrombotic microangiopathy and HUS induced by EHEC Shiga toxins in these preclinical models, and benefits or risks of complement inhibition should be studied further for this infection.

  14. Shiga toxin 1 interaction with enterocytes causes apical protein mistargeting through the depletion of intracellular galectin-3

    Energy Technology Data Exchange (ETDEWEB)

    Laiko, Marina; Murtazina, Rakhilya; Malyukova, Irina [Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205 (United States); Zhu, Chengru; Boedeker, Edgar C. [Department of Medicine, University of New Mexico School of Medicine, Albuquerque, NM 87131 (United States); Gutsal, Oksana [Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205 (United States); O' Malley, Robert; Cole, Robert N. [Department of Biochemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205 (United States); Tarr, Phillip I. [Department of Pediatrics, Washington University School of Medicine, St. Louis, MO 63110 (United States); Murray, Karen F. [Department of Pediatrics, Children' s Hospital and Regional Medical Center, Seattle, WA 98105 (United States); Kane, Anne [The Tufts New England Medical Center, Boston, MA 02111 (United States); Donowitz, Mark [Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205 (United States); Kovbasnjuk, Olga, E-mail: okovbas1@jhmi.edu [Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205 (United States)

    2010-02-15

    Shiga toxins (Stx) 1 and 2 are responsible for intestinal and systemic sequelae of infection by enterohemorrhagic Escherichia coli (EHEC). However, the mechanisms through which enterocytes are damaged remain unclear. While secondary damage from ischemia and inflammation are postulated mechanisms for all intestinal effects, little evidence excludes roles for more primary toxin effects on intestinal epithelial cells. We now document direct pathologic effects of Stx on intestinal epithelial cells. We study a well-characterized rabbit model of EHEC infection, intestinal tissue and stool samples from EHEC-infected patients, and T84 intestinal epithelial cells treated with Stx1. Toxin uptake by intestinal epithelial cells in vitro and in vivo causes galectin-3 depletion from enterocytes by increasing the apical galectin-3 secretion. This Shiga toxin-mediated galectin-3 depletion impairs trafficking of several brush border structural proteins and transporters, including villin, dipeptidyl peptidase IV, and the sodium-proton exchanger 2, a major colonic sodium absorptive protein. The mistargeting of proteins responsible for the absorptive function might be a key event in Stx1-induced diarrhea. These observations provide new evidence that human enterocytes are directly damaged by Stx1. Conceivably, depletion of galectin-3 from enterocytes and subsequent apical protein mistargeting might even provide a means whereby other pathogens might alter intestinal epithelial absorption and produce diarrhea.

  15. Colonization of Enteroaggregative Escherichia coli and Shiga toxin-producing Escherichia coli in chickens and humans in southern Vietnam

    NARCIS (Netherlands)

    Trung, Nguyen Vinh; Nhung, Hoang Ngoc; Carrique-Mas, Juan J.; Mai, Ho Huynh; Tuyen, Ha Thanh; Campbell, James; Nhung, Nguyen Thi; van Minh, Pham; Wagenaar, Jaap A.; Mai, Nguyen Thi Nhu; Hieu, Thai Quoc; Schultsz, Constance; Hoa, Ngo Thi

    2016-01-01

    Enteroaggregative (EAEC) and Shiga-toxin producing Escherichia coli (STEC) are a major cause of diarrhea worldwide. E. coli carrying both virulence factors characteristic for EAEC and STEC and producing extended-spectrum beta-lactamase caused severe and protracted disease during an outbreak of E.

  16. Classification of non-O157 shiga toxin-producing escherichia coli(STEC) serotypes with hyperspectral microscope imaging

    Science.gov (United States)

    Non-O157 Shiga toxin-producing Escherichia coli (STEC) strains such as O26, O45, O103, O111, O121 and O145 are recognized as serious outbreak to cause human illness due to their toxicity. A conventional microbiological method for cell counting is laborious and needs long time for the results. Since ...

  17. Protozoan Predation of Escherichia coli O157:H7 Is Unaffected by the Carriage of Shiga Toxin-Encoding Bacteriophages.

    Directory of Open Access Journals (Sweden)

    Carrie E Schmidt

    Full Text Available Escherichia coli O157:H7 is a food-borne bacterium that causes hemorrhagic diarrhea and hemolytic uremic syndrome in humans. While cattle are a known source of E. coli O157:H7 exposure resulting in human infection, environmental reservoirs may also be important sources of infection for both cattle and humans. Bacteriophage-encoded Shiga toxins (Stx carried by E. coli O157:H7 may provide a selective advantage for survival of these bacteria in the environment, possibly through their toxic effects on grazing protozoa. To determine Stx effects on protozoan grazing, we co-cultured Paramecium caudatum, a common ciliate protozoon in cattle water sources, with multiple strains of Shiga-toxigenic E. coli O157:H7 and non-Shiga toxigenic cattle commensal E. coli. Over three days at ambient laboratory temperature, P. caudatum consistently reduced both E. coli O157:H7 and non-Shiga toxigenic E. coli populations by 1-3 log cfu. Furthermore, a wild-type strain of Shiga-toxigenic E. coli O157:H7 (EDL933 and isogenic mutants lacking the A subunit of Stx 2a, the entire Stx 2a-encoding bacteriophage, and/or the entire Stx 1-encoding bacteriophage were grazed with similar efficacy by both P. caudatum and Tetrahymena pyriformis (another ciliate protozoon. Therefore, our data provided no evidence of a protective effect of either Stx or the products of other bacteriophage genes on protozoan predation of E. coli. Further research is necessary to determine if the grazing activity of naturally-occurring protozoa in cattle water troughs can serve to decrease cattle exposure to E. coli O157:H7 and other Shiga-toxigenic E. coli.

  18. Shiga toxin-producing Escherichia coli in meat: a preliminary simulation study on detection capabilities for three sampling methods

    Science.gov (United States)

    The objective of this simulation study is to determine which sampling method (Cozzini core sampler, core drill shaving, and N-60 surface excision) will better detect Shiga Toxin-producing Escherichia coli (STEC) at varying levels of contamination when present in the meat. 1000 simulated experiments...

  19. Mathematical modeling of growth of non-O157 Shiga Toxin-producing Escherichia coli in raw ground beef

    Science.gov (United States)

    The objective of this study was to investigate the growth of Shiga toxin-producing Escherichia coli (STEC, including serogroups O45, O103, O111, O121, and O145) in raw ground beef and to develop mathematical models to describe the bacterial growth under different temperature conditions. Three prima...

  20. Inactivation of a diverse set of shiga toxin-producing Escherichia coli in ground beef by high pressure processing

    Science.gov (United States)

    Shiga Toxin-Producing Escherichia coli (STEC) are frequently implicated in foodborne illness outbreaks and recalls of ground beef. In this study we determined the High Pressure Processing (HPP) D-10 value (the processing conditions needed to reduce the microbial population by 1 log) of 39 individua...

  1. Induction of Shiga Toxin-Encoding Prophage by Abiotic Environmental Stress in Food.

    Science.gov (United States)

    Fang, Yuan; Mercer, Ryan G; McMullen, Lynn M; Gänzle, Michael G

    2017-10-01

    The prophage-encoded Shiga toxin is a major virulence factor in Stx-producing Escherichia coli (STEC). Toxin production and phage production are linked and occur after induction of the RecA-dependent SOS response. However, food-related stress and Stx-prophage induction have not been studied at the single-cell level. This study investigated the effects of abiotic environmental stress on stx expression by single-cell quantification of gene expression in STEC O104:H4 Δ stx2 :: gfp :: amp r In addition, the effect of stress on production of phage particles was determined. The lethality of stressors, including heat, HCl, lactic acid, hydrogen peroxide, and high hydrostatic pressure, was selected to reduce cell counts by 1 to 2 log CFU/ml. The integrity of the bacterial membrane after exposure to stress was measured by propidium iodide (PI). The fluorescent signals of green fluorescent protein (GFP) and PI were quantified by flow cytometry. The mechanism of prophage induction by stress was evaluated by relative gene expression of recA and cell morphology. Acid (pH stress were additionally assessed. H 2 O 2 and mitomycin C induced expression of the prophage and activated a SOS response. In contrast, HCl and lactic acid induced the Stx-prophage but not the SOS response. The lifestyle of STEC exposes the organism to intestinal and extraintestinal environments that impose oxidative and acid stress. A more thorough understanding of the influence of food processing-related stressors on Stx-prophage expression thus facilitates control of STEC in food systems by minimizing prophage induction during food production and storage. Copyright © 2017 American Society for Microbiology.

  2. Distribution and detection of Shiga toxin-producing Escherichia coli (STEC) during an industrial grinding process of beef trim

    Science.gov (United States)

    During the grinding and packaging processes, it is important to understand how Shiga toxin-producing Escherichia coli (STEC) would be distributed and how well it could be detected in beef trim. This study is important because it shows what would happen if contaminated meat is allowed into a commerc...

  3. Shiga Toxin—A Model for Glycolipid-Dependent and Lectin-Driven Endocytosis

    Directory of Open Access Journals (Sweden)

    Ludger Johannes

    2017-10-01

    Full Text Available The cellular entry of the bacterial Shiga toxin and the related verotoxins has been scrutinized in quite some detail. This is due to their importance as a threat to human health. At the same time, the study of Shiga toxin has allowed the discovery of novel molecular mechanisms that also apply to the intracellular trafficking of endogenous proteins at the plasma membrane and in the endosomal system. In this review, the individual steps that lead to Shiga toxin uptake into cells will first be presented from a purely mechanistic perspective. Membrane-biological concepts will be highlighted that are often still poorly explored, such as fluctuation force-driven clustering, clathrin-independent membrane curvature generation, friction-driven scission, and retrograde sorting on early endosomes. It will then be explored whether and how these also apply to other pathogens, pathogenic factors, and cellular proteins. The molecular nature of Shiga toxin as a carbohydrate-binding protein and that of its cellular receptor as a glycosylated raft lipid will be an underlying theme in this discussion. It will thereby be illustrated how the study of Shiga toxin has led to the proposal of the GlycoLipid-Lectin (GL-Lect hypothesis on the generation of endocytic pits in processes of clathrin-independent endocytosis.

  4. Different Effects of Six Antibiotics and Ten Traditional Chinese Medicines on Shiga Toxin Expression by Escherichia coli O157:H7

    Directory of Open Access Journals (Sweden)

    Mei Ling Chen

    2013-01-01

    Full Text Available This study compared the effects of ten types of traditional Chinese medicines (TCMs and six different antibiotics on E. coli O157:H7 Shiga toxin gene (stx2 mRNA expression level based on real-time PCR and the expression level of Stx toxin using an ELISA quantitative assay. We also compared their effects on the induction of the SOS response. The results clearly indicated that all ten TCMs had negative results in the SOS response induction test, while most TCMs did not increase the levels of stx2 mRNA and the Stx toxin. Some TCMs did increase the mRNA levels of the stx2 gene and the Stx toxin level, but their increases were much lower than those caused by antibiotics. With the exception of cefotaxime, the six antibiotics increased the Stx toxin level and increased the stx2 gene mRNA level. With the exceptions of cefotaxime and tetracycline, the antibiotics increased the SOS induction response. These results suggest that TCMs may have advantages compared with antibiotics, when treating E. coli O157:H7; TCMs did not greatly increase Stx toxin production and release.

  5. N-chlorotaurine, a long-lived oxidant produced by human leukocytes, inactivates Shiga toxin of enterohemorrhagic Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Christian Eitzinger

    Full Text Available N-chlorotaurine (NCT, the main representative of long-lived oxidants produced by granulocytes and monocytes, is known to exert broad-spectrum microbicidal activity. Here we show that NCT directly inactivates Shiga toxin 2 (Stx2, used as a model toxin secreted by enterohemorrhagic Escherichia coli (EHEC. Bacterial growth and Stx2 production were both inhibited by 2 mM NCT. The cytotoxic effect of Stx2 on Vero cells was removed by ≥5.5 mM NCT. Confocal microscopy and FACS analyses showed that the binding of Stx2 to human kidney glomerular endothelial cells was inhibited, and no NCT-treated Stx2 entered the cytosol. Mass spectrometry displayed oxidation of thio groups and aromatic amino acids of Stx2 by NCT. Therefore, long-lived oxidants may act as powerful tools of innate immunity against soluble virulence factors of pathogens. Moreover, inactivation of virulence factors may contribute to therapeutic success of NCT and novel analogs, which are in development as topical antiinfectives.

  6. Carbon nanoparticles in lateral flow methods to detect genes encoding virulence factors of Shiga toxin-producing Escherichia coli

    NARCIS (Netherlands)

    Noguera, P.; Posthuma-Trumpie, G.A.; Tuil, van M.; Wal, van der F.J.; Boer, de A.; Moers, A.P.H.A.; Amerongen, van A.

    2011-01-01

    The use of carbon nanoparticles is shown for the detection and identification of different Shiga toxin-producing Escherichia coli virulence factors (vt1, vt2, eae and ehxA) and a 16S control (specific for E. coli) based on the use of lateral flow strips (nucleic acid lateral flow immunoassay,

  7. Validation of the Applied Biosystems RapidFinder Shiga Toxin-Producing E. coli (STEC) Detection Workflow.

    Science.gov (United States)

    Cloke, Jonathan; Matheny, Sharon; Swimley, Michelle; Tebbs, Robert; Burrell, Angelia; Flannery, Jonathan; Bastin, Benjamin; Bird, Patrick; Benzinger, M Joseph; Crowley, Erin; Agin, James; Goins, David; Salfinger, Yvonne; Brodsky, Michael; Fernandez, Maria Cristina

    2016-11-01

    The Applied Biosystems™ RapidFinder™ STEC Detection Workflow (Thermo Fisher Scientific) is a complete protocol for the rapid qualitative detection of Escherichia coli (E. coli) O157:H7 and the "Big 6" non-O157 Shiga-like toxin-producing E. coli (STEC) serotypes (defined as serogroups: O26, O45, O103, O111, O121, and O145). The RapidFinder STEC Detection Workflow makes use of either the automated preparation of PCR-ready DNA using the Applied Biosystems PrepSEQ™ Nucleic Acid Extraction Kit in conjunction with the Applied Biosystems MagMAX™ Express 96-well magnetic particle processor or the Applied Biosystems PrepSEQ Rapid Spin kit for manual preparation of PCR-ready DNA. Two separate assays comprise the RapidFinder STEC Detection Workflow, the Applied Biosystems RapidFinder STEC Screening Assay and the Applied Biosystems RapidFinder STEC Confirmation Assay. The RapidFinder STEC Screening Assay includes primers and probes to detect the presence of stx1 (Shiga toxin 1), stx2 (Shiga toxin 2), eae (intimin), and E. coli O157 gene targets. The RapidFinder STEC Confirmation Assay includes primers and probes for the "Big 6" non-O157 STEC and E. coli O157:H7. The use of these two assays in tandem allows a user to detect accurately the presence of the "Big 6" STECs and E. coli O157:H7. The performance of the RapidFinder STEC Detection Workflow was evaluated in a method comparison study, in inclusivity and exclusivity studies, and in a robustness evaluation. The assays were compared to the U.S. Department of Agriculture (USDA), Food Safety and Inspection Service (FSIS) Microbiology Laboratory Guidebook (MLG) 5.09: Detection, Isolation and Identification of Escherichia coli O157:H7 from Meat Products and Carcass and Environmental Sponges for raw ground beef (73% lean) and USDA/FSIS-MLG 5B.05: Detection, Isolation and Identification of Escherichia coli non-O157:H7 from Meat Products and Carcass and Environmental Sponges for raw beef trim. No statistically significant

  8. Epithelial and Mesenchymal Cells in the Bovine Colonic Mucosa Differ in Their Responsiveness to Escherichia coli Shiga Toxin 1

    Science.gov (United States)

    Cells in the depth of the crypts in the bovine colon express CD77 molecules that potentially act as receptors for Shiga toxins (Stx). The implication of this finding for the intestinal colonization 25 of cattle with human pathogenic Stx-producing Escherichia coli (STEC) remains undefined. We used f...

  9. Carbon nanoparticles as detection labels in antibody microarrays. Detection of genes encoding virulence factors in Shiga toxin-producing Escherichia coli.

    NARCIS (Netherlands)

    Noguera, P.S.; Posthuma-Trumpie, G.A.; Tuil, Van M.; Wal, van der F.J.; Boer, De A.; Moers, A.P.H.A.; Amerongen, Van A.

    2011-01-01

    The present study demonstrates that carbon nanoparticles (CNPs) can be used as labels in microarrays. CNPs were used in nucleic acid microarray immunoassays (NAMIAs) for the detection of different Shiga toxin-producing Escherichia coli (STEC) virulence factors: four genes specific for STEC (vt1,

  10. The Role of the Exo-Xis Region in Oxidative Stress-Mediated Induction of Shiga Toxin-Converting Prophages

    Directory of Open Access Journals (Sweden)

    Katarzyna Licznerska

    2016-01-01

    Full Text Available Previous studies indicated that these genetic elements could be involved in the regulation of lysogenization and prophage induction processes. The effects were dramatic in Shiga toxin-converting phage Φ24B after treatment with oxidative stress-inducing agent, hydrogen peroxide, while they were less pronounced in bacteriophage λ and in both phages irradiated with UV. The hydrogen peroxide-caused prophage induction was found to be RecA-dependent. Importantly, in hydrogen peroxide-treated E. coli cells lysogenic for either λ or Φ24B, deletion of the exo-xis region resulted in a significant decrease in the levels of expression of the S.O.S. regulon genes. Moreover, under these conditions, a dramatic decrease in the levels of expression of phage genes crucial for lytic development (particularly xis, exo, N, cro, O, Q, and R could be observed in Φ24B-, but not in λ-bearing cells. We conclude that genes located in the exo-xis region are necessary for efficient expression of both host S.O.S regulon in lysogenic bacteria and regulatory genes of Shiga toxin-converting bacteriophage Φ24B.

  11. Occurrence and characterization of shiga toxin-producing Escherichia coli in raw meat, raw milk, and street vended juices in Bangladesh

    NARCIS (Netherlands)

    Islam, M.A.; Mondol, A.S.; Azmi, I.J.; Boer, de E.; Beumer, R.R.; Zwietering, M.H.; Heuvelink, A.E.; Talukder, K.A.

    2010-01-01

    The major objective of this study was to investigate the prevalence of Shiga toxin (Stx)–producing Escherichia coli (STEC) in different types of food samples and to compare their genetic relatedness with STEC strains previously isolated from animal sources in Bangladesh. We investigated a total of

  12. Risk Factors for Salmonella, Shiga Toxin-Producing Escherichia coli and Campylobacter Occurrence in Primary Production of Leafy Greens and Strawberries

    Directory of Open Access Journals (Sweden)

    Siele Ceuppens

    2015-08-01

    Full Text Available The microbiological sanitary quality and safety of leafy greens and strawberries were assessed in the primary production in Belgium, Brazil, Egypt, Norway and Spain by enumeration of Escherichia coli and detection of Salmonella, Shiga toxin-producing E. coli (STEC and Campylobacter. Water samples were more prone to containing pathogens (54 positives out of 950 analyses than soil (16/1186 and produce on the field (18/977 for leafy greens and 5/402 for strawberries. The prevalence of pathogens also varied markedly according to the sampling region. Flooding of fields increased the risk considerably, with odds ratio (OR 10.9 for Salmonella and 7.0 for STEC. A significant association between elevated numbers of generic E. coli and detection of pathogens (OR of 2.3 for STEC and 2.7 for Salmonella was established. Generic E. coli was found to be a suitable index organism for Salmonella and STEC, but to a lesser extent for Campylobacter. Guidelines on frequency of sampling and threshold values for E. coli in irrigation water may differ from region to region.

  13. From farm to table: follow-up of Shiga toxin-producing Escherichia coli throughout the pork production chain in Argentina

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    Rocío eColello

    2016-02-01

    Full Text Available Pigs are important reservoirs of Shiga toxin-producing Escherichia coli (STEC. The entrance of these strains into the food chain implies a risk to consumers because of the severity of hemolytic uremic syndrome. This study reports the prevalence and characterization of Shiga toxin-producing Escherichia coli (STEC throughout the pork production chain. From 764 samples, 31 (4.05% were stx positive by PCR screening. At farms, 2.86% of samples were stx positive; at slaughter, 4.08% of carcasses were stx positive and at boning rooms, 6% of samples were stx positive. These percentages decreased in pork meat ready for sale at sales markets (4.59%. From positive samples, 50 isolates could be characterized. At farms 37.5% of the isolates carried stx1/stx2 genes, 37.5% possessed stx2e and 25%, carried only stx2. At slaughter we detected 50% of isolates positive for stx2, 33% for stx2e and 16% for stx1/stx2. At boning rooms 59% of the isolates carried stx1/stx2, 14% stx2e and 5% stx1/stx2/stx2e. At retail markets 66% of isolates were positive for stx2, 17% stx2e and 17% stx1/stx2. For the other virulence factors, ehxA and saa were not detected and eae gene was detected in 12% of the isolates. Concerning putative adhesins, agn43 was detected in 72%, ehaA in 26%, aida in 8% and iha in 6% of isolates. The strains were typed into 14 E. coli O groups (O1, O2, O8, O15, O20, O35, O69, O78, O91, O121, O138, O142, O157, O180 and ten H groups (H9, H10, H16, H21, H26, H29, H30, H32, H45, H46. This study reports the prevalence and characterization of STEC strains through the chain pork suggesting the vertical transmission. STEC contamination originates in the farms and is transferred from pigs to carcasses in the slaughter process and increase in meat pork at boning rooms and sales markets. These results highlight the need to implement an integrated STEC control system based on good management practices on the farm and critical control point systems in the food chain.

  14. Mouse in Vivo Neutralization of Escherichia coli Shiga Toxin 2 with Monoclonal Antibodies

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    Larry H. Stanker

    2013-10-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC food contaminations pose serious health concerns, and have been the subject of massive food recalls. STEC has been identified as the major cause of the life-threatening complication of hemolytic uremic syndrome (HUS. Besides supportive care, there currently are no therapeutics available. The use of antibiotics for combating pathogenic E. coli is not recommended because they have been shown to stimulate toxin production. Clearing Stx2 from the circulation could potentially lessen disease severity. In this study, we tested the in vivo neutralization of Stx2 in mice using monoclonal antibodies (mAbs. We measured the biologic half-life of Stx2 in mice and determined the distribution phase or t1/2 α to be 3 min and the clearance phase or t1/2 β to be 40 min. Neutralizing mAbs were capable of clearing Stx2 completely from intoxicated mouse blood within minutes. We also examined the persistence of these mAbs over time and showed that complete protection could be passively conferred to mice 4 weeks before exposure to Stx2. The advent of better diagnositic methods and the availability of a greater arsenal of therapeutic mAbs against Stx2 would greatly enhance treatment outcomes of life threatening E. coli infections.

  15. Enteroaggregative, Shiga Toxin-Producing Escherichia coli O111:H2 Associated with an Outbreak of Hemolytic-Uremic Syndrome

    Science.gov (United States)

    Morabito, Stefano; Karch, Helge; Mariani-Kurkdjian, Patrizia; Schmidt, Herbert; Minelli, Fabio; Bingen, Edouard; Caprioli, Alfredo

    1998-01-01

    Shiga toxin-producing Escherichia coli O111:H2 strains from an outbreak of hemolytic-uremic syndrome showed aggregative adhesion to HEp-2 cells and harbored large plasmids which hybridized with the enteroaggregative E. coli probe PCVD432. These strains present a novel combination of virulence factors and might be as pathogenic to humans as the classic enterohemorrhagic E. coli. PMID:9508328

  16. Effect of high pressure impact on the survival of Shiga Toxin-producing Escherichia coli ('Big Six' and 0157) in ground beef

    Science.gov (United States)

    High pressure processing (HPP) is a safe and effective technology for improving food safety while maintaining food quality attributes. Non-O157:H7 Shiga Toxin-producing Escherichia coli (STEC) have been increasingly implicated in foodborne illness outbreaks and recalls, and the USDA Food Safety Ins...

  17. Phenotypic and genotypic characteristics of Shiga toxin-producing Escherichia coli isolated from surface waters and sediments in a Canadian urban-agricultural landscape

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    Stephanie eNadya

    2016-04-01

    Full Text Available A hydrophobic grid membrane filtration – Shiga toxin immunoblot method was used to examine the prevalence of Shiga toxin-producing Escherichia coli (STEC in four watersheds located in the Lower Mainland of British Columbia, Canada, a region characterized by rapid urbanization and intensive agricultural activity. STEC were recovered from 21.6, 23.2, 19.5 and 9.2 % of surface water samples collected monthly from five sites in each watershed over a period of one year. Overall prevalence was subject to seasonal variation however, ranging between 13.3 % during fall months and 34.3 % during winter months. STEC were also recovered from 23.8 % of sediment samples collected in one randomly selected site. One hundred distinct STEC isolates distributed among 29 definitive and 4 ambiguous or indeterminate serotypes were recovered from water and sediments, including isolates from Canadian priority serogroups O157 (3, O26 (4, O103 (5 and O111 (7. Forty seven isolates were further characterized by analysis of whole genome sequences to detect Shiga toxin gene (stx 1 and stx 2, intimin gene (eaeA allelic variants and acquired virulence factors. These analyses collectively showed that surface waters from the region support highly diverse STEC populations that include strains with virulence factors commonly associated with human pathotypes. The present work served to characterize the microbiological hazard implied by STEC to support future assessments of risks to public health arising from non-agricultural and agricultural uses of surface water resources in the region.

  18. Mathematical modeling and numerical analysis of the growth of Non-O157 shiga toxin-producing Escherichia coli in spinach leaves

    Science.gov (United States)

    This study was conducted to investigate the growth of non-O157 Shiga toxin-producing Escherichia coli (STEC) in spinach leaves and to develop kinetic models to describe the bacterial growth. Six serogroups of non-O157 STEC, including O26, O45, O103, O111, O121, and O145, were used in the growth stu...

  19. Upregulation of Shiga toxin receptor CD77/Gb3 and interleukin-1β expression in the brain of EHEC patients with hemolytic uremic syndrome and neurologic symptoms.

    Science.gov (United States)

    Hagel, Christian; Krasemann, Susanne; Löffler, Judith; Püschel, Klaus; Magnus, Tim; Glatzel, Markus

    2015-03-01

    In 2011, a large outbreak of Shiga toxin-producing enterohemorrhagic Escherichia coli (EHEC) infections occurred in northern Germany, which mainly affected adults. Out of 3842 patients, 104 experienced a complicated course comprising hemolytic uremic syndrome and neurological complications, including cognitive impairment, aphasia, seizures and coma. T2 hyperintensities on magnet resonance imaging (MRI) bilateral in the thalami and in the dorsal pons were found suggestive of a metabolic toxic effect. Five of the 104 patients died because of toxic heart failure. In the present study, the post-mortem neuropathological findings of the five EHEC patients are described. Histological investigation of 13 brain regions (frontal, temporal, occipital cortex, corpora mammillaria, thalamus, frontal operculum, corona radiata, gyrus angularis, pons, medulla oblongata, cerebellar vermis and cerebellar hemisphere) showed no thrombosis, ischemic changes or fresh infarctions. Further, no changes were found in electron microscopy. In comparison with five age-matched controls, slightly increased activation of microglia and a higher neuronal expression of interleukin-1β and of Shiga toxin receptor CD77/globotriaosylceramide 3 was observed. The findings were confirmed by Western blot analyses. It is suggested that CD77/globotriaosylceramide upregulation may be a consequence to Shiga toxin exposure, whereas increased interleukin-1β expression may point to activation of inflammatory cascades. © 2014 International Society of Neuropathology.

  20. Subtype-specific suppression of Shiga toxin 2 released from Escherichia coli upon exposure to protein synthesis inhibitors

    DEFF Research Database (Denmark)

    Pedersen, Malene Gantzhorn; Hansen, Claus; Riise, Erik

    2008-01-01

    Shiga toxins (Stx) are important virulence factors in the pathogenesis of severe disease including hemolytic-uremic syndrome, caused by Stx-producing Escherichia coli (STEC). STEC strains increase the release of Stx in vitro following the addition of fluoroquinolones, whereas protein synthesis...... inhibitors previously have been reported to suppress the release of Stx. The amount of Stx released from wild-type STEC strains incubated with protein synthesis inhibitors was examined by a Vero cell cytotoxicity assay. The amounts released were compared to the Stx type (Stx1 or Stx2) and additionally...... to the individual subtypes and toxin variants of Stx2. In general, Stx2 release was suppressed significantly upon exposure to protein synthesis inhibitors at MICs, which was not observed in the case of Stx1. Also, the average amount of different Stx2 toxin variants released was suppressed to various levels ranging...

  1. Molecular basis of differential B-pentamer stability of Shiga toxins 1 and 2.

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    Deborah G Conrady

    2010-12-01

    Full Text Available Escherichia coli strain O157:H7 is a major cause of food poisoning that can result in severe diarrhea and, in some cases, renal failure. The pathogenesis of E. coli O157:H7 is in large part due to the production of Shiga toxin (Stx, an AB(5 toxin that consists of a ribosomal RNA-cleaving A-subunit surrounded by a pentamer of receptor-binding B subunits. There are two major isoforms, Stx1 and Stx2, which differ dramatically in potency despite having 57% sequence identity. Animal studies and epidemiological studies show Stx2 is associated with more severe disease. Although the molecular basis of this difference is unknown, data suggest it is associated with the B-subunit. Mass spectrometry studies have suggested differential B-pentamer stability between Stx1 and Stx2. We have examined the relative stability of the B-pentamers in solution. Analytical ultracentrifugation using purified B-subunits demonstrates that Stx2B, the more deadly isoform, shows decreased pentamer stability compared to Stx1B (EC(50 = 2.3 µM vs. EC(50 = 0.043 µM for Stx1B. X-ray crystal structures of Stx1B and Stx2B identified a glutamine in Stx2 (versus leucine in Stx1 within the otherwise strongly hydrophobic interface between B-subunits. Interchanging these residues switches the stability phenotype of the B-pentamers of Stx1 and Stx2, as demonstrated by analytical ultracentrifugation and circular dichroism. These studies demonstrate a profound difference in stability of the B-pentamers in Stx1 and Stx2, illustrate the mechanistic basis for this differential stability, and provide novel reagents to test the basis for differential pathogenicity of these toxins.

  2. Detection and characterization of Shiga toxin-producing Escherichia coli from seagulls.

    Science.gov (United States)

    Makino, S; Kobori, H; Asakura, H; Watarai, M; Shirahata, T; Ikeda, T; Takeshi, K; Tsukamoto, T

    2000-08-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) strains isolated from a seagull in Japan were examined. A total of 50 faecal samples was collected on a harbour bank in Hokkaido, Japan, in July 1998. Two different STEC strains, whose serotypes were O136:H16 and O153:H-, were isolated from the same individual by PCR screening; both of them were confirmed by ELISA and Vero cell cytotoxicity assay to be producing active Stx2 and Stx1, respectively. They harboured large plasmids, but did not carry the haemolysin or eaeA genes of STEC O157:H7. Based on their plasmid profiles, antibiotic resistance patterns, pulsed-field gel electrophoresis analysis (PFGE), and the stx genes sequences, the isolates were different. Phylogenic analysis of the deduced Stx amino acid sequences demonstrated that the Stx toxins of seagull-origin STEC were closely associated with those of the human-origin, but not those of other animal-origin STEC. In addition, Stx2phi-K7 phage purified from O136 STEC resembled Stx2phi-II from human-origin O157:H7, and was able to convert non-toxigenic E. coli to STEC. These results suggest that birds may be one of the important carriers in terms of the distribution of STEC.

  3. Characterization of free lytic bacteriophages isolated from compost against O145 Shiga toxin-producing Escherichia coli (STEC) as a potential biocontrol agent

    Science.gov (United States)

    Shiga toxin-producing E. coli (STEC), one of the most prevalent foodborne pathogens, are notorious for hemolytic uremic syndrome (HUS) and causing high mortality among children and the elder population after infection. Besides O157 STEC, non-O157 STEC—particularly serogroup O145—is commonly associat...

  4. Influence of RNase E deficiency on the production of stx2-bearing phages and Shiga toxin in an RNase E-inducible strain of enterohaemorrhagic Escherichia coli (EHEC) O157:H7.

    Science.gov (United States)

    Thuraisamy, Thujitha; Lodato, Patricia B

    2018-05-01

    In enterohaemorrhagic Escherichia coli (EHEC), stx1 or stx2 genes encode Shiga toxin (Stx1 or Stx2, respectively) and are carried by prophages. The production and release of both stx phages and toxin occur upon initiation of the phage lytic cycle. Phages can further disseminate stx genes by infecting naïve bacteria in the intestine. Here, the effect of RNase E deficiency on these two virulence traits was investigated. Cultures of the EHEC strains TEA028-rne containing low versus normal RNase E levels or the parental strain (TEA028) were treated with mitomycin C (MMC) to induce the phage lytic cycle. Phages and Stx2 titres were quantified by the double-agar assay and the receptor ELISA technique, respectively. RNase E deficiency in MMC-treated cells significantly reduced the yield of infectious stx2 phages. Delayed cell lysis and the appearance of encapsidated phage DNA copies suggest a slow onset of the lytic cycle. However, these observations do not entirely explain the decrease of phage yields. stx1 phages were not detected under normal or deficient RNase E levels. After an initial delay, high levels of toxin were finally produced in MMC-treated cultures. RNase E scarcity reduces stx2 phage production but not toxin. Normal concentrations of RNase E are likely required for correct phage morphogenesis. Our future work will address the mechanism of RNase E action on phage morphogenesis.

  5. In vivo screening platform for shiga toxin-producing Escherichia coli (STEC using Caenorhabditis elegans as a model.

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    Su-Bin Hwang

    Full Text Available Shiga toxin-producing Escherichia coli (STEC strains are the main cause of bacillary dysentery, although STEC strains generally induce milder disease symptoms compared to Shigella species. This study aimed to determine the virulence of STEC using the nematode Caenorhabditis elegans as a model host. Worm killing, fertility and bacterial colonisation assays were performed to examine the potential difference in the virulence of STEC strains compared to that of the control E. coli OP50 strains on which worms were fed. A statistically significant difference in the survival rates of C. elegans was observed in that the STEC strains caused death in 8-10 days and the E. coli OP50 strains caused death in 15 days. STEC strains severely reduced the fertility of the worms. The intestinal load of bacteria in the adult stage nematodes harbouring the E. coli OP50 strains was found to be 3.5 log CFU mL-1. In contrast, the STEC strains E15, E18 and E22 harboured 4.1, 4.2 and 4.7 log CFU ml-1 per nematode, respectively. The heat-killed STEC strains significantly increased the longevity of the worms compared to the non-heated STEC strains. In addition, PCR-based genomic profiling of shiga toxin genes, viz., stx1 and stx2, identified in selected STEC strains revealed that these toxins may be associated with the virulence of the STEC strains. This study demonstrated that C. elegans is an effective model to examine and compare the pathogenicity and virulence variation of STEC strains to that of E. coli OP50 strains.

  6. Anti-bacterial effect of essential oil from Xanthium strumarium against shiga toxin-producing Escherichia coli.

    Science.gov (United States)

    Sharifi-Rad, J; Soufi, L; Ayatollahi, S A M; Iriti, M; Sharifi-Rad, M; Varoni, E M; Shahri, F; Esposito, S; Kuhestani, K; Sharifi-Rad, M

    2016-09-19

    Shiga toxin-producing Escherichia coli (STEC) serotype O157:H7 is one of the most important human pathogenic microorganisms, which can cause life-threatening infections. Xanthium strumarium L. is a plant with anti-bacterial activity against gram-negative and gram-positive bacteria. This study aims to demonstrate in vitro efficacy of the essential oil (EO) extracted from Xanthium strumarium L. against E. coli O157:H7. Using the agar test diffusion, the effect of Xanthium strumarium L. EO (5, 10, 15, 30, 60, and 120 mg/mL) was verified at each of the four different growth phases of E. coli O157:H7. Cell counts of viable cells and colony forming unit (CFU) were determined at regular time points using Breed's method and colony counting method, respectively. No viable cell was detectable after the 1 hour-exposure to X. strumarium EO at 30, 60, and 120 mg/mL concentrations. No bacterial colony was formed after 1 h until the end of the incubation period at 24 h. At lower concentrations, the number of bacteria cells decreased and colonies could be observed only after incubation. At the exponential phase, the EO at 15 mg/mL was only bacteriostatic, while from 30 mg/mL started to be bactericidal. X. strumarium EO antibacterial activity against Shiga toxin-producing E. coli O157:H7 is dependent on EO concentration and physiological state of the microorganisms tested. The best inhibitory activity was achieved during the late exponential and the stationary phases.

  7. Prevalence of sorbitol non-fermenting Shiga toxin-producing Escherichia coli in Black Bengal goats on smallholdings.

    Science.gov (United States)

    Das Gupta, M; Das, A; Islam, M Z; Biswas, P K

    2016-09-01

    A cross-sectional survey was carried out in Bangladesh with the sampling of 514 Black Bengal goats on smallholdings to determine the presence of sorbitol non-fermenting (SNF) Shiga toxin-producing E. coli (STEC). Swab samples collected from the recto-anal junction were plated onto cefixime and potassium tellurite added sorbitol MacConkey (CT-SMAC) agar, a selective medium for STEC O157 serogroup, where this serogroup and other SNF STEC produce colourless colonies. The SNF E. coli (SNF EC) isolates obtained from the survey were investigated by PCR for the presence of Shiga toxin-producing genes, stx1 and stx2, and two other virulence genes, eae and hlyA that code for adherence factor (intimin protein) and pore-forming cytolysin, respectively. The SNF EC isolates were also assessed for the presence of the rfbO157 gene to verify their identity to O157 serogroup. The results revealed that the proportions of goats carrying SNF EC isolates and stx1 and stx2 genes were 6·2% (32/514) [95% confidence interval (CI) 4·4-8·7)], 1·2% (95% CI 0·5-2·6) and 1·2% (95% CI 0·5-2·6), respectively. All the SNF STEC tested negative for rfbO157, hlyA and eae genes. The risk for transmission of STEC from Black Bengal goats to humans is low.

  8. The effect of deep frying or conventional oven cooking on inactivation of Shiga toxin-producing cells of Escherichia coli (STEC) in meatballs

    Science.gov (United States)

    We investigated the effects deep frying or oven cooking on inactivation of Shiga toxin-producing cells of Escherichia coli (STEC) in meatballs. A finely-ground veal and/or a beef-pork-veal mixture were inoculated (ca. 7.0 log CFU/g) with an eight-strain, genetically-marked cocktail of rifampicin-res...

  9. Assessment of Consumer Exposure to Salmonella spp., Campylobacter spp., and Shiga Toxin-Producing Escherichia coli in Meat Products at Retail in the City of Sao Paulo, Brazil.

    Science.gov (United States)

    Ristori, Christiane Asturiano; Rowlands, Ruth Estela Gravato; Martins, Cecília Geraldes; Barbosa, Maria Luisa; Dos Santos, Luis Fernando; Jakabi, Miyoko; de Melo Franco, Bernadette Dora Gombossy

    2017-08-01

    Meat products may be vehicles of bacterial pathogens to humans, and Salmonella spp., Campylobacter spp., and Shiga toxin-producing Escherichia coli (STEC) are the most relevant. The aim of this study was to generate data on prevalence of these three pathogens in 552 samples of meat products (hot dogs, pork sausages, raw ground beef, and raw chicken legs) sold at retail in the city of Sao Paulo, Brazil. Salmonella spp. was detected in 5.8% (32/552) of samples, comprising pork sausages 62.5% (20/32) and chicken legs 37.5% (12/32). The counts of Salmonella spp. were low, ranging from Salmonella Typhimurium (28.1%), Salmonella I 4,[5],12:i:- (15.6%), Salmonella Enteritidis (12.5%), Salmonella Derby, and Salmonella Brandenburg (9.4%). Campylobacter spp. was detected in 33 samples (6.0%), comprising chicken legs (82%) and ground beef (18%). All samples were negative for STEC. These results suggest that meat products when subjected to inadequate cooking and/or cross-contamination with other products ready for consumption can lead to occurrence of outbreaks, highlighting the risks associated with them.

  10. Strain-Level Discrimination of Shiga Toxin-Producing Escherichia coli in Spinach Using Metagenomic Sequencing.

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    Susan R Leonard

    Full Text Available Consumption of fresh bagged spinach contaminated with Shiga toxin-producing Escherichia coli (STEC has led to severe illness and death; however current culture-based methods to detect foodborne STEC are time consuming. Since not all STEC strains are considered pathogenic to humans, it is crucial to incorporate virulence characterization of STEC in the detection method. In this study, we assess the comprehensiveness of utilizing a shotgun metagenomics approach for detection and strain-level identification by spiking spinach with a variety of genomically disparate STEC strains at a low contamination level of 0.1 CFU/g. Molecular serotyping, virulence gene characterization, microbial community analysis, and E. coli core gene single nucleotide polymorphism (SNP analysis were performed on metagenomic sequence data from enriched samples. It was determined from bacterial community analysis that E. coli, which was classified at the phylogroup level, was a major component of the population in most samples. However, in over half the samples, molecular serotyping revealed the presence of indigenous E. coli which also contributed to the percent abundance of E. coli. Despite the presence of additional E. coli strains, the serotype and virulence genes of the spiked STEC, including correct Shiga toxin subtype, were detected in 94% of the samples with a total number of reads per sample averaging 2.4 million. Variation in STEC abundance and/or detection was observed in replicate spiked samples, indicating an effect from the indigenous microbiota during enrichment. SNP analysis of the metagenomic data correctly placed the spiked STEC in a phylogeny of related strains in cases where the indigenous E. coli did not predominate in the enriched sample. Also, for these samples, our analysis demonstrates that strain-level phylogenetic resolution is possible using shotgun metagenomic data for determining the genomic relatedness of a contaminating STEC strain to other

  11. Comparative Genomic and Phylogenetic Analysis of a Shiga Toxin Producing Shigella sonnei (STSS Strain

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    Domonkos Sváb

    2017-05-01

    Full Text Available Shigella strains are important agents of bacillary dysentery, and in recent years Shigella sonnei has emerged as the leading cause of shigellosis in industrialized and rapidly developing countries. More recently, several S. sonnei and Shigella flexneri strains producing Shiga toxin (Stx have been reported from sporadic cases and from an outbreak in America. In the present study we aimed to shed light on the evolution of a recently identified Shiga toxin producing S. sonnei (STSS isolated in Europe. Here we report the first completely assembled whole genome sequence of a multidrug resistant (MDR Stx-producing S. sonnei (STSS clinical strain and reveal its phylogenetic relations. STSS 75/02 proved to be resistant to ampicillin, streptomycin, tetracycline, chloramphenicol, thrimetoprim, and sulfomethoxazol. The genome of STSS 75/02 contains a 4,891,717 nt chromosome and seven plasmids including the 214 kb invasion plasmid (pInv harboring type III secretion system genes and associated effectors. The chromosome harbors 23 prophage regions including the Stx1 converting prophage. The genome carries all virulence determinants necessary for an enteroinvasive lifestyle, as well as the Stx1 encoding gene cluster within an earlier described inducible converting prophage. In silico SNP genotyping of the assembled genome as well as 438 complete or draft S. sonnei genomes downloaded from NCBI GenBank revealed that S. sonnei 75/02 belongs to the more recently diverged global MDR lineage (IIIc. Targeted screening of 1131 next-generation sequencing projects taken from NCBI Short Read Archive of confirms that only a few S. sonnei isolates are Stx positive. Our results suggest that the acquisition of Stx phages could have occurred in different environments as independent events and that multiple horizontal transfers are responsible for the appearance of Stx phages in S. sonnei strains.

  12. Pectic oligosaccharide structure-function relationships: prebiotics, inhibitors of Escherichia coli O157:H7 adhesion and reduction of Shiga toxin cytotoxicity in HT29 cells

    Science.gov (United States)

    Shiga toxin (Stx)-producing, food-contaminating Escherichia coli (STEC) is a major health concern. Plant-derived pectin and pectic-oligosaccharides (POS) that are abundant in biomass have been considered as prebiotics and for the protection of humans from Stx intoxication. Five structurally differ...

  13. Incidence of Shiga toxin-producing Escherichia coli strains in beef, pork, chicken, deer, boar, bison, and rabbit retail meat.

    Science.gov (United States)

    Magwedere, Kudakwashe; Dang, Huu Anh; Mills, Edward W; Cutter, Catherine N; Roberts, Elisabeth L; DeBroy, Chitrita

    2013-03-01

    The objective of the current study was to determine the incidence of contamination by the top 7 Shiga toxin-producing Escherichia coli (STEC) O-groups, responsible for the majority of E. coli infections in human beings, in retail meat from different animal species. Samples from ground beef (n = 51), ground pork (n = 16), ground chicken (n = 16), and game meat (deer, wild boar, bison, and rabbit; n = 55) were collected from retail vendors for the detection of 7 STEC O-groups (O26, O45, O103, O111, O121, O145, and O157). Meat samples were tested by using a multiplex polymerase chain reaction assay targeting the wzx gene of O antigen gene clusters of the 7 STEC O-groups. The positive samples were further tested for Shiga toxin genes (stx1 and stx2). Out of a total of 83 ground beef, pork, and chicken samples, 17 (20%) carried O121, 9 (10%) carried O45, 8 (9%) carried O157, 3 (3%) carried O103, and 1 (1%) carried O145. None of the samples were positive for O26, O111, or the stx gene. All 3 white-tailed deer samples (100%) were positive for O45, O103, or both, 2 (10%) out of 20 red deer samples exhibited the presence of O103, and all 3 bison samples were contaminated with either O121, O145, or O157. One sample from ground deer, contaminated with E. coli O45, carried the stx1 gene. This preliminary investigation illustrates the importance of microbiological testing of pathogens in meat products, as well as the recognized need for increased surveillance and research on foodborne pathogens.

  14. Variation in the Distribution of Putative Virulence and Colonization Factors in Shiga Toxin-Producing Escherichia coli Isolated from Different Categories of Cattle

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    Analía I. Etcheverría

    2017-04-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC are pathogens of significant public health concern. Several studies have confirmed that cattle are the main reservoir of STEC in Argentina and other countries. Although Shiga toxins represent the primary virulence factors of STEC, the adherence and colonization of the gut are also important in the pathogenesis of the bacteria. The aim of this study was to analyze and to compare the presence of putative virulence factors codified in plasmid -katP, espP, subA, stcE- and adhesins involved in colonization of cattle -efa1, iha- in 255 native STEC strains isolated from different categories of cattle from different production systems. The most prevalent gene in all strains was espP, and the less prevalent was stcE. katP was highly detected in strains isolated from young and rearing calves (33.3%, while subA was predominant in those isolated from adults (71.21%. Strains from young calves showed the highest percentage of efa1 (72.46%, while iha showed a high distribution in strains from rearing calves and adults (87.04 and 98.48% respectively. It was observed that espP and iha were widely distributed throughout all strains, whereas katP, stcE, and efa1 were more associated with the presence of eae and subA with the eae-negative strains. A great proportion of eae-negative strains were isolated from adults -dairy and grazing farms- and from rearing calves -dairy and feedlot-, while mostly of the eae-positive strains were isolated from dairy young calves. Data exposed indicate a correlation between the category of the animal and the production systems with the presence or absence of several genes implicated in adherence and virulence of STEC.

  15. Immunoconcentration of Shiga toxin-producing Escherichia coli O157 from animal faeces and raw meats by using Dynabeads anti-E. coli O157 and the VIDAS system

    NARCIS (Netherlands)

    Islam, M.A.; Heuvelink, A.E.; Talukder, K.A.; Boer, de E.

    2006-01-01

    To identify the reservoirs and routes of transmission of Shiga toxin-producing Escherichia coli (STEC) O157, sensitive detection and isolation methods are necessary. The sensitivity of traditional culture methods can be improved significantly by the inclusion of an immunoconcentration step,

  16. Survival and expression of acid resistance genes in Shiga toxin-producing Escherichia coli acid adapted in pineapple juice and exposed to synthetic gastric fluid

    Science.gov (United States)

    Aims: The aim of this research was to examine relative transcriptional expression of acid resistance (AR) genes, rpoS, gadA and adiA, in O157:H7 and non-O157 Shiga toxin-producing Escherichia coli (STEC) serotypes after adaptation to pineapple juice (PJ) and subsequently to determine survival with e...

  17. Isolation of bacteriophages against non-O157 and O157 Shiga toxin-producing Escherichia coli (STEC) from composting of non-fecal materials and the potential impact on produce safety

    Science.gov (United States)

    Composting is a complex process to produce fertilizers used to improve crop yields. A complete composting process usually confers bactericidal effect due to change of temperature and pH However, some produce outbreaks associated with Shiga toxin-producing E. coli (STEC) contamination were linked to ...

  18. In Vivo Tumor Targeting by the B-Subunit of Shiga Toxin

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    Thomas Viel

    2008-11-01

    Full Text Available Delivery of drugs to the appropriate target cells would improve efficacy and reduce potential side effects. The nontoxic B-subunit of the intestinal pathogen-produced Shiga toxin (STxB binds specifically to the glycosphingolipid Gb3, overex-pressed in membranes of certain tumor cells, and enters these cells through the retrograde pathway. Therefore, STxB binding to Gb3 receptors may be useful for cell-specific vectorization or imaging purposes. Here we labeled STxB with a fluorophore to evaluate its potential as an in vivo cell-specific targeting reagent in two different models of human colorectal carcinoma. Fluorescent STxB was administered systemically to xenografted nude mice, and its biodistribution was studied by optical imaging. The use of fluorescent STxB allowed the combination of the macroscopic observations with analyses at the cellular level using confocal microscopy. After administration, the fluorescent STxB was slowly eliminated by renal excretion. However, it accumulated in the tumor area. Furthermore, STxB was demonstrated to enter the Gb3-expressing tumoral cells, as well as the epithelial cells of the neovascularization and the monocytes and macrophages surrounding the xenografts.

  19. Molecular Characterization of Shiga Toxin-Producing Escherichia coli Isolated from Ruminant and Donkey Raw Milk Samples and Traditional Dairy Products in Iran

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    Hassan Momtaz

    2012-01-01

    Full Text Available The aims of the current study were to detect the virulence factors and antibiotic resistance of Shiga toxin-producing E. coli, in animal milk and dairy products in Iran. After E. coli dentification with culture method, PCR assay were developed for detection of pathogenic genes, serotypes and antibiotic resistance genes of E. coli. Results showed that out of 719 samples, 102 (14.18% were confirmed to be positive for E. coli and out of 102 positive samples, 17.64% were O26 and 13.72% were O157 and 1.96% were O91 and 1.96% were O145 serotypes. Totally, the prevalence of stx1 and papA genes were the highest while the prevalence of sfaS and fyuA were the lowest in the positive samples. PCR results showed that tetA, tetB were the highest (64.70% and aac(3-IV were the lowest (27.45% antibiotic resistant genes in E. coli positive samples. Our study indicated that the isolated E. coli trains in these regions had a highest antibiotic resistance to tetracycline (58.82% and the lowest to nitrofurantoin (3.92%. tetA gene and E. coli O157 serotype had highest and aac(3-IV gene, and E. coli O145 serotype had a lowest frequency rates of antibiotics resistance genes, in the region.

  20. Detection of E.Coli Strains Containing Shiga Toxin (Stx1/2 Gene in Diarrheal Specimens from Children Less than 5 Years Old by PCR Technique and Study of the Patterns of Antibiotic Resistance

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    MR Pourmand

    2009-10-01

    Full Text Available Introduction: Shiga toxin- producing Escherichia coli (STEC is an emerging bacterial pathogen in developing countries that causes several diseases such as diarrhea, hemorrhagic colitis (HC and hemolytic uremic syndrome (HUS, particularly in children. Aim of the research was detection of STEC in diarrheal specimens from under 5 year olds and study of the patterns of antibiotic resistance of these strains. Methods: In the study,300 fecal samples were collected from children with diarrhea referring to Ali Asghar Hospital. E.coli species were isolated by standard bacteriological and biochemical tests. Presence of shiga toxin genes (stx1/2 was investigated by PCR technique (Qiagen. Antibiogram test for strains containing the toxin gene was performed using 16 different antibiotic discs (MAST by disc diffusion agar (Kirby-Bauer method. Results: From 39 E.coli isolates, 9(23.1% strains were detected by PCR to contain stx1/2 gene. One strain was resistant to all 16 antibiotics. All the STEC strains were sensitive to meropenem (MRP, imipenem (IMI, gentamycin (GEN and nitrofurantoin (NI. 4(44.44% strains showed multi-drug resistant pattern. All these 4strains were resistant to cotrimoxazole(SxT. Also, 6(66.66% strains were resistant to at least one antibiotic. Conclusion: In Iran, shiga toxin- producing Escherichia coli (STEC may be a commonly bacterial pathogen causing diarrhea, particularly in children. Therefore, we should use new techniques for investigation of these strains. Increase in number of emerging and new strains that could be resistant to classic antibiotics such as cotrimoxazole may be foreseen. It is suggested that antibiotics prescription programs in treatment of diarrhea causing E.coli strains be updated.

  1. Specific egg yolk immunoglobulin as a new preventive approach for Shiga-toxin-mediated diseases.

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    Paola Neri

    Full Text Available Shiga toxins (Stxs are involved in the development of severe systemic complications associated with enterohemorrhagic Escherichia coli (EHEC infection. Various neutralizing agents against Stxs are under investigation for management of EHEC infection. In this study, we immunized chickens with formalin-inactivated Stx-1 or Stx-2, and obtained immunoglobulin Y (IgY from the egg yolk. Anti-Stx-1 IgY and anti-Stx-2 IgY recognized the corresponding Stx A subunit and polymeric but not monomeric B subunit. Anti-Stx-1 IgY and anti-Stx-2 IgY suppressed the cytotoxicity of Stx-1 and Stx-2 to HeLa 229 cells, without cross-suppressive activity. The suppressive activity of these IgY was abrogated by pre-incubation with the corresponding recombinant B subunit, which suggests that the antibodies directed to the polymeric B subunits were predominantly involved in the suppression. In vivo, the intraperitoneal or intravenous administration of these IgY rescued mice from death caused by intraperitoneal injection of the corresponding toxin at a lethal dose. Moreover, oral administration of anti-Stx-2 IgY reduced the mortality of mice infected intestinally with EHEC O157:H7. Our results therefore suggest that anti-Stx IgY antibodies may be considered as preventive agents for Stx-mediated diseases in EHEC infection.

  2. Bad Phages in Good Bacteria: Role of the Mysterious orf63 of λ and Shiga Toxin-Converting Φ24B Bacteriophages

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    Aleksandra Dydecka

    2017-08-01

    Full Text Available Lambdoid bacteriophages form a group of viruses that shares a common schema of genome organization and lifecycle. Some of them can play crucial roles in creating the pathogenic profiles of Escherichia coli strains. For example, Shiga toxin-producing E. coli (STEC acquired stx genes, encoding Shiga toxins, via lambdoid prophages (Stx phages. The results obtained so far present the evidence for the relation between the exo-xis region of the phage genome and lambdoid phage development, however molecular mechanisms of activities of the exo-xis genes' products are still unknown. In view of this, we decided to determine the influence of the uncharacterized open reading frame orf63 of the exo-xis region on lambdoid phages development using recombinant prophages, λ and Stx phage Φ24B. We have demonstrated that orf63 codes for a folded protein, thus, it is a functional gene. NMR spectroscopy and analytical gel filtration were used to extend this observation further. From backbone chemical shifts, Orf63 is oligomeric in solution, likely a trimer and consistent with its small size (63 aa., is comprised of two helices, likely intertwined to form the oligomer. We observed that the deletion of phage orf63 does not impair the intracellular lambdoid phage lytic development, however delays the time and decreases the efficiency of prophage induction and in consequence results in increased survival of E. coli during phage lytic development. Additionally, the deletion of phage orf63 negatively influences expression of the major phage genes and open reading frames from the exo-xis region during prophage induction with hydrogen peroxide. We conclude, that lambdoid phage orf63 may have specific functions in the regulation of lambdoid phages development, especially at the stage of the lysis vs. lysogenization decision. Besides, orf63 probably participates in the regulation of the level of expression of essential phage genes and open reading frames from the exo

  3. Shiga toxin-producing Escherichia coli distribution and characterization in a pasture-based cow-calf production system.

    Science.gov (United States)

    Baltasar, Patrícia; Milton, Stewart; Swecker, William; Elvinger, François; Ponder, Monica

    2014-05-01

    Shiga toxin-producing Escherichia coli (STEC) strains are commonly found in cattle gastrointestinal tracts. In this study, prevalence and distribution of E. coli virulence genes (stx1, stx2, hlyA, and eaeA) were assessed in a cow-calf pasture-based production system. Angus cows (n = 90) and their calves (n = 90) were kept in three on-farm locations, and fecal samples were collected at three consecutive times (July, August, and September 2011). After enrichment of samples, stx1, stx2, eaeA, and hlyA were amplified and detected with a multiplex PCR (mPCR) assay. Fecal samples positive for stx genes were obtained from 93.3% (84 of 90) of dams and 95.6% (86 of 90) of calves at one or more sampling times. Age class (dam or calf), spatial distribution of cattle (farm locations B, H, K), and sampling time influenced prevalence and distribution of virulence genes in the herd. From 293 stx-positive fecal samples, 744 E. coli colonies were isolated. Virulence patterns of isolates were determined through mPCR assay: stx1 was present in 41.9% (312 of 744) of the isolates, stx2 in 6.5% (48 of 744), eaeA in 4.2% (31 of 744), and hlyA in 2.4% (18 of 744). Prevalence of non-O157 STEC was high among the isolates: 33.8% (112 of 331) were STEC O121, 3.6% (12 of 331) were STEC O103, and 1.8% (6 of 331) were STEC O113. One isolate (0.3%) was identified as STEC O157. Repetitive element sequence-based PCR (rep-PCR) fingerprinting was used to study genetic diversity of stx-positive E. coli isolates. Overall, rep-PCR fingerprints were highly similar, supporting the hypothesis that strains are transmitted between animals but not necessarily from a dam to its calf. Highly similar STEC isolates were obtained at each sampling time, but isolates obtained from dams were more diverse than those from calves, suggesting that strain differences in transference may exist. Understanding the transfer of E. coli from environmental and animal sources to calves may aid in developing intervention

  4. Detection and isolation of the "Top 7" Shiga toxin-producing Escherichia coli in ground beef: comparison of the Rapidfinder kits to the USDA microbiology laboratory guidebook method

    Science.gov (United States)

    Shiga toxin-producing E. coli (STEC) O157:H7 and serogroups O26, O45, O103, O111, O121, and O145 are often referred to as the “top 7” STEC, and these have been declared as adulterants in beef by the USDA Food Safety and Inspection Service (FSIS). The aim of this work was to compare the methods des...

  5. Proteomic View of Interactions of Shiga Toxin-Producing Escherichia coli with the Intestinal Environment in Gnotobiotic Piglets.

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    Rembert Pieper

    Full Text Available Shiga toxin (Stx-producing Escherichia coli cause severe intestinal infections involving colonization of epithelial Peyer's patches and formation of attachment/effacement (A/E lesions. These lesions trigger leukocyte infiltration followed by inflammation and intestinal hemorrhage. Systems biology, which explores the crosstalk of Stx-producing Escherichia coli with the in vivo host environment, may elucidate novel molecular pathogenesis aspects.Enterohemorrhagic E. coli strain 86-24 produces Shiga toxin-2 and belongs to the serotype O157:H7. Bacterial cells were scrapped from stationary phase cultures (the in vitro condition and used to infect gnotobiotic piglets via intestinal lavage. Bacterial cells isolated from the piglets' guts constituted the in vivo condition. Cell lysates were subjected to quantitative 2D gel and shotgun proteomic analyses, revealing metabolic shifts towards anaerobic energy generation, changes in carbon utilization, phosphate and ammonia starvation, and high activity of a glutamate decarboxylase acid resistance system in vivo. Increased abundance of pyridine nucleotide transhydrogenase (PntA and PntB suggested in vivo shortage of intracellular NADPH. Abundance changes of proteins implicated in lipopolysaccharide biosynthesis (LpxC, ArnA, the predicted acyltransferase L7029 and outer membrane (OM assembly (LptD, MlaA, MlaC suggested bacterial cell surface modulation in response to activated host defenses. Indeed, there was evidence for interactions of innate immunity-associated proteins secreted into the intestines (GP340, REG3-γ, resistin, lithostathine, and trefoil factor 3 with the bacterial cell envelope.Proteomic analysis afforded insights into system-wide adaptations of strain 86-24 to a hostile intestinal milieu, including responses to limited nutrients and cofactor supplies, intracellular acidification, and reactive nitrogen and oxygen species-mediated stress. Protein and lipopolysaccharide compositions of the OM

  6. Antibody Microarray for E. coli O157:H7 and Shiga Toxin in Microtiter Plates

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    Andrew G. Gehring

    2015-12-01

    Full Text Available Antibody microarray is a powerful analytical technique because of its inherent ability to simultaneously discriminate and measure numerous analytes, therefore making the technique conducive to both the multiplexed detection and identification of bacterial analytes (i.e., whole cells, as well as associated metabolites and/or toxins. We developed a sandwich fluorescent immunoassay combined with a high-throughput, multiwell plate microarray detection format. Inexpensive polystyrene plates were employed containing passively adsorbed, array-printed capture antibodies. During sample reaction, centrifugation was the only strategy found to significantly improve capture, and hence detection, of bacteria (pathogenic Escherichia coli O157:H7 to planar capture surfaces containing printed antibodies. Whereas several other sample incubation techniques (e.g., static vs. agitation had minimal effect. Immobilized bacteria were labeled with a red-orange-fluorescent dye (Alexa Fluor 555 conjugated antibody to allow for quantitative detection of the captured bacteria with a laser scanner. Shiga toxin 1 (Stx1 could be simultaneously detected along with the cells, but none of the agitation techniques employed during incubation improved detection of the relatively small biomolecule. Under optimal conditions, the assay had demonstrated limits of detection of ~5.8 × 105 cells/mL and 110 ng/mL for E. coli O157:H7 and Stx1, respectively, in a ~75 min total assay time.

  7. Antibody Microarray for E. coli O157:H7 and Shiga Toxin in Microtiter Plates.

    Science.gov (United States)

    Gehring, Andrew G; Brewster, Jeffrey D; He, Yiping; Irwin, Peter L; Paoli, George C; Simons, Tawana; Tu, Shu-I; Uknalis, Joseph

    2015-12-04

    Antibody microarray is a powerful analytical technique because of its inherent ability to simultaneously discriminate and measure numerous analytes, therefore making the technique conducive to both the multiplexed detection and identification of bacterial analytes (i.e., whole cells, as well as associated metabolites and/or toxins). We developed a sandwich fluorescent immunoassay combined with a high-throughput, multiwell plate microarray detection format. Inexpensive polystyrene plates were employed containing passively adsorbed, array-printed capture antibodies. During sample reaction, centrifugation was the only strategy found to significantly improve capture, and hence detection, of bacteria (pathogenic Escherichia coli O157:H7) to planar capture surfaces containing printed antibodies. Whereas several other sample incubation techniques (e.g., static vs. agitation) had minimal effect. Immobilized bacteria were labeled with a red-orange-fluorescent dye (Alexa Fluor 555) conjugated antibody to allow for quantitative detection of the captured bacteria with a laser scanner. Shiga toxin 1 (Stx1) could be simultaneously detected along with the cells, but none of the agitation techniques employed during incubation improved detection of the relatively small biomolecule. Under optimal conditions, the assay had demonstrated limits of detection of ~5.8 × 10⁵ cells/mL and 110 ng/mL for E. coli O157:H7 and Stx1, respectively, in a ~75 min total assay time.

  8. Prevalence and Characterization of Shiga Toxin-Producing Escherichia coli Isolated from Slaughtered Qurban Animal in Jakarta Province

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    Siti Gusti Ningrum

    2016-08-01

    Full Text Available This study was conducted to investigate the presence of shiga toxin producing Escherichia coli (STEC and the possibility of carrying rfbE gene and H7 flagellar on meat, liver, and stool samples collected from Jakarta Province of Indonesia. A total of 51 samples collected from meat, liver, and stool of slaughtered cattle from qurban festival were tested using conventional culture and multiplex PCR methods. STEC non O157 were detected in meat (5.3% and stool (8.3% with one isolate from stool carried H7 flagellar. However, all isolates were lacking of rfbE gene. In antimicrobial susceptibility tests, the STEC isolates showed antibiotic resistance to erythromycin and oxacillin. Overall, the result shows that meat and liver of this origin activity represents a potential risk to human health.

  9. Prevalence and characterization of Listeria monocytogenes, Salmonella and Shiga toxin-producing Escherichia coli isolated from small Mexican retail markets of queso fresco.

    Science.gov (United States)

    Soto Beltran, Marcela; Gerba, Charles P; Porto Fett, Anna; Luchansky, John B; Chaidez, Cristobal

    2015-01-01

    Queso fresco (QF) is a handmade cheese consumed and produced in Latin America. In Mexico, QF production is associated with a microbiological risk. The aim of the study was to determine the incidence and characterization of Listeria monocytogenes, Salmonella spp., and Shiga toxin-producing Escherichia coli (STEC) in QF from retail markets of the north-western State of Sinaloa, Mexico, and to assess the effect of physicochemical parameters on Listeria presence. A total of 75 QF samples were obtained. L. monocytogenes, E. coli, and coliforms were detected in 9.3, 94, and 100%, respectively. Salmonella was not detected. STEC isolates showed virulence genes. Microbial loads were above the maximum values recommended by the Official Mexican Standards. Physicochemical parameters such as water activity (aw), moisture content, pH, and salinity played a role in Listeria prevalence in QF. Rigorous control in QF made in Culiacan, Mexico is needed to reduce the risk of foodborne pathogens.

  10. Occurrence and characterization of Shiga toxin-producing Escherichia coli O157:H7 and other non-sorbitol-fermenting E. coli in cattle and humans in urban areas of Morogoro, Tanzania

    DEFF Research Database (Denmark)

    Lupindu, Athumani M; Olsen, John Elmerdahl; Ngowi, Helena A

    2014-01-01

    Escherichia coli strains such as Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli, enterotoxigenic, attaching, and effacing E. coli, and enteroinvasive E. coli cause diarrhea in humans. Although other serotypes exist, the most commonly reported STEC in outbreaks is O157:H7. A cross-...

  11. Induction of Shiga toxin-converting prophage in Escherichia coli by high hydrostatic pressure.

    Science.gov (United States)

    Aertsen, Abram; Faster, David; Michiels, Chris W

    2005-03-01

    Since high hydrostatic pressure is becoming increasingly important in modern food preservation, its potential effects on microorganisms need to be thoroughly investigated. In this context, mild pressures (pressures. In this report, we extend this observation to lambdoid Shiga toxin (Stx)-converting bacteriophages in MG1655, which constitute an important virulence trait in Stx-producing E. coli strains (STEC). The window of pressures capable of inducing Stx phages correlated well with the window of bacterial survival. When pressure treatments were conducted in whole milk, which is known to promote bacterial survival, Stx phage induction could be observed at up to 250 MPa in E. coli MG1655 and at up to 300 MPa in a pressure-resistant mutant of this strain. In addition, we found that the intrinsic pressure resistance of two types of Stx phages was very different, with one type surviving relatively well treatments of up to 400 MPa for 15 min at 20 degrees C. Interestingly, and in contrast to UV irradiation or mitomycin C treatment, pressure was not able to induce Stx prophage or an SOS response in several natural Stx-producing STEC isolates.

  12. Comparative evaluation of the Ridascreen Verotoxin enzyme immunoassay for detection of Shiga-toxin producing strains of Escherichia coli (STEC) from food and other sources.

    Science.gov (United States)

    Beutin, L; Steinrück, H; Krause, G; Steege, K; Haby, S; Hultsch, G; Appel, B

    2007-03-01

    To evaluate the suitability of the commercially distributed Ridascreen Verotoxin enzyme immunoassay (EIA) for detection of known genetic types of the Vero (Shiga) toxins 1 (Stx1) and 2 (Stx2) families and to determine its relative sensitivity and specificity. The Ridascreen-EIA was compared with the Vero cell assay, a P(1)-glycoprotein receptor EIA and with stx gene-specific PCs for detection of Stx with 43 Shiga toxin-producing strains of Escherichia coli (STEC) reference strains and with 241 test strains. The Ridascreen-EIA detects strains producing Stx1 and variants Stx1c and Stx1d, as well as Stx2 and variants Stx2d1, Stx2d2, Stx2e, Stx2d, Stx2-O118 (Stx2d-ount), Stx2-NV206, Stx2f and Stx2g. The assay showed a relative sensitivity of 95.7% and a relative specificity of 98.7%. Some of the Stx2-O118-, Stx2e- and Stx2g-producing STEC were not detected with the Ridascreen-EIA probably because of low amount of toxin produced by these strains. The Ridascreen-EIA is able to detect all known types of Stx and is applicable for routine screening of bacterial isolates owing to its high specificity. It is less applicable for testing samples where low amounts of Stx are expected, such as mixed cultures and certain Stx2 variants. This study presents a first comprehensive evaluation of the Ridascreen-EIA, a rapid standardized STEC screening test for routine diagnostic laboratories. Data are presented on the type of the spectrum of Stx that are detected with this immunoassay and its advantages and limits for practical use.

  13. Shiga toxin-producing Escherichia coli isolated from chicken meat in Iran: serogroups, virulence factors, and antimicrobial resistance properties.

    Science.gov (United States)

    Momtaz, Hassan; Jamshidi, Alireza

    2013-05-01

    The aim of the current study was to determine the virulence factors, serogroups, and antibiotic resistance properties of Shiga toxin-producing Escherichia coli isolated from chicken meat samples. A total of 422 chicken meat samples were collected from 5 townships of Iran. Specimens were immediately transferred to the laboratory in a cooler with an ice pack. Samples were cultured, and the positive culture samples were analyzed by PCR assays. Finally, the antimicrobial susceptibility test was performed using the disk diffusion method in Mueller-Hinton agar. According to the results, out of 422 samples, 146 (34.59%) were confirmed to be E. coli positive and among E. coli-positive samples, 51 (34.93%) and 31 (21.23%) were from attaching and effacing E. coli (AEEC) and enterohemorrhagic E. coli (EHEC) subgroups, respectively. All of the EHEC-positive samples had all stx1, eaeA, and ehly virulence genes, whereas only 5 (9.80%) of AEEC subgroup had all stx1, stx2, and eaeA genes. As the data revealed, O157 was the most prevalent and O111 was the least prevalent strains in the Shiga toxin-producing E. coli (STEC) population. Among STEC strains, sulI and blaSHV had the highest and lowest incidence rate, respectively. There was a high resistance to tetracycline (76.82%), followed by chloramphenicol (73.17%) and nitrofurantoin (63.41%), but there was low resistance to cephalotine (7.31%) antibiotics in isolated strains. Results shows that the PCR technique has a high performance for detection of serogroups, virulence genes, and antibiotic resistance genes in STEC strains. This study is the first prevalence report of detection of virulence genes, serogroups, and antibiotic resistance properties of STEC strains isolated from chicken meat samples in Iran. Based on the results, chicken meat is one of the main sources of STEC strains and its virulence factors in Iran, so an accurate meat inspection would reduce disease outbreaks.

  14. Growth of Escherichia coli O157:H7, Non-O157 Shiga Toxin-Producing Escherichia coli , and Salmonella in Water and Hydroponic Fertilizer Solutions.

    Science.gov (United States)

    Shaw, Angela; Helterbran, Kara; Evans, Michael R; Currey, Christopher

    2016-12-01

    The desire for local, fresh produce year round is driving the growth of hydroponic growing systems in the United States. Many food crops, such as leafy greens and culinary herbs, grown within hydroponics systems have their root systems submerged in recirculating nutrient-dense fertilizer solutions from planting through harvest. If a foodborne pathogen were introduced into this water system, the risk of contamination to the entire crop would be high. Hence, this study was designed to determine whether Escherichia coli O157:H7, non-O157 Shiga toxin-producing E. coli , and Salmonella were able to survive and reproduce in two common hydroponic fertilizer solutions and in water or whether the bacteria would be killed or suppressed by the fertilizer solutions. All the pathogens grew by 1 to 6 log CFU/ml over a 24-h period, depending on the solution. E. coli O157:H7 reached higher levels in the fertilizer solution with plants (3.12 log CFU/ml), whereas non-O157 Shiga toxin-producing E. coli and Salmonella reached higher levels in the fertilizer solution without plants (1.36 to 3.77 log CFU/ml). The foodborne pathogens evaluated here survived for 24 h in the fertilizer solution, and populations grew more rapidly in these solutions than in plain water. Therefore, human pathogens entering the fertilizer solution tanks in hydroponic systems would be expected to rapidly propagate and spread throughout the system and potentially contaminate the entire crop.

  15. Prevalence and Characterization of Shiga Toxin-Producing Escherichia coli in Swine Feces Recovered in the National Animal Health Monitoring System's Swine 2000 Study

    Science.gov (United States)

    Fratamico, Pina M.; Bagi, Lori K.; Bush, Eric J.; Solow, Barbara T.

    2004-01-01

    A study was conducted to determine the prevalence of Shiga toxin-producing Escherichia coli (STEC) in swine feces in the United States as part of the National Animal Health Monitoring System's Swine 2000 study. Fecal samples collected from swine operations from 13 of the top 17 swine-producing states were tested for the presence of STEC. After enrichment of swine fecal samples in tryptic soy broth, the samples were tested for the presence of stx1 and stx2 by use of the TaqMan E. coli STX1 and STX2 PCR assays. Enrichments of samples positive for stx1 and/or stx2 were plated, and colony hybridization was performed using digoxigenin-labeled probes complementary to the stx1 and stx2 genes. Positive colonies were picked and confirmed by PCR for the presence of the stx1, stx2, or stx2e genes, and the isolates were serotyped. Out of 687 fecal samples tested using the TaqMan assays, 70% (484 of 687) were positive for Shiga toxin genes, and 54% (370 of 687), 64% (436 of 687), and 38% (261 of 687) were positive for stx1, stx2, and both toxin genes, respectively. Out of 219 isolates that were characterized, 29 (13%) produced stx1, 14 (6%) produced stx2, and 176 (80%) produced stx2e. Twenty-three fecal samples contained at least two STEC strains that had different serotypes but that had the same toxin genes or included a strain that possessed stx1 in addition to a strain that possessed stx2 or stx2e. The STEC isolates belonged to various serogroups, including O2, O5, O7, O8, O9, OX10, O11, O15, OX18, O20, O57, O65, O68, O69, O78, O91, O96, O100, O101, O120, O121, O152, O159, O160, O163, and O untypeable. It is noteworthy that no isolates of serogroup O157 were recovered. Results of this study indicate that swine in the United States harbor STEC that can potentially cause human illness. PMID:15574914

  16. DNA microarray-based assessment of virulence potential of Shiga toxin gene-carrying Escherichia coli O104:H7 isolated from feedlot cattle feces.

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    Pragathi B Shridhar

    Full Text Available Escherichia coli O104:H4, a hybrid pathotype reported in a large 2011 foodborne outbreak in Germany, has not been detected in cattle feces. However, cattle harbor and shed in the feces other O104 serotypes, particularly O104:H7, which has been associated with sporadic cases of diarrhea in humans. The objective of our study was to assess the virulence potential of Shiga toxin-producing E. coli (STEC O104:H7 isolated from feces of feedlot cattle using DNA microarray. Six strains of STEC O104:H7 isolated from cattle feces were analyzed using FDA-E. coli Identification (ECID DNA microarray to determine their virulence profiles and compare them to the human strains (clinical of O104:H7, STEC O104:H4 (German outbreak strain, and O104:H21 (milk-associated Montana outbreak strain. Scatter plots were generated from the array data to visualize the gene-level differences between bovine and human O104 strains, and Pearson correlation coefficients (r were determined. Splits tree was generated to analyze relatedness between the strains. All O104:H7 strains, both bovine and human, similar to O104:H4 and O104:H21 outbreak strains were negative for intimin (eae. The bovine strains were positive for Shiga toxin 1 subtype c (stx1c, enterohemolysin (ehxA, tellurite resistance gene (terD, IrgA homolog protein (iha, type 1 fimbriae (fimH, and negative for genes that code for effector proteins of type III secretory system. The six cattle O104 strains were closely related (r = 0.86-0.98 to each other, except for a few differences in phage related and non-annotated genes. One of the human clinical O104:H7 strains (2011C-3665 was more closely related to the bovine O104:H7 strains (r = 0.81-0.85 than the other four human clinical O104:H7 strains (r = 0.75-0.79. Montana outbreak strain (O104:H21 was more closely related to four of the human clinical O104:H7 strains than the bovine O104:H7 strains. None of the bovine E. coli O104 strains carried genes characteristic of E

  17. Ouabain Protects Human Renal Cells against the Cytotoxic Effects of Shiga Toxin Type 2 and Subtilase Cytotoxin

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    María M. Amaral

    2017-07-01

    Full Text Available Hemolytic uremic syndrome (HUS is one of the most common causes of acute renal failure in children. The majority of cases are associated with Shiga toxin (Stx-producing Escherichia coli (STEC. In Argentina, HUS is endemic and presents the highest incidence rate in the world. STEC strains expressing Stx type 2 (Stx2 are responsible for the most severe cases of this pathology. Subtilase cytotoxin (SubAB is another STEC virulence factor that may contribute to HUS pathogenesis. To date, neither a licensed vaccine nor effective therapy for HUS is available for humans. Considering that Ouabain (OUA may prevent the apoptosis process, in this study we evaluated if OUA is able to avoid the damage caused by Stx2 and SubAB on human glomerular endothelial cells (HGEC and the human proximal tubule epithelial cell (HK-2 line. HGEC and HK-2 were pretreated with OUA and then incubated with the toxins. OUA protected the HGEC viability from Stx2 and SubAB cytotoxic effects, and also prevented the HK-2 viability from Stx2 effects. The protective action of OUA on HGEC and HK-2 was associated with a decrease in apoptosis and an increase in cell proliferation. Our data provide evidence that OUA could be considered as a therapeutic strategy to avoid the renal damage that precedes HUS.

  18. The new allelic variant of the subtilase cytotoxin (subAB2) is common among Shiga toxin-producing Escherichia coli strains from large game animals and their meat and meat products.

    Science.gov (United States)

    Sánchez, Sergio; Díaz-Sánchez, Sandra; Martínez, Remigio; Llorente, María Teresa; Herrera-León, Silvia; Vidal, Dolors

    2013-10-25

    Subtilase cytotoxin (SubAB) is an AB5 toxin produced by Shiga toxin (Stx)-producing Escherichia coli (STEC) strains usually lacking the eae gene product intimin. Two allelic variants of SubAB encoding genes have been described: subAB1, located on a plasmid, and subAB2, located on a pathogenicity island (PAI) together with tia gene. While subAB1 has been reported to be more frequent among bovine strains, subAB2 has been mainly associated with strains from small ruminants. We investigated the presence of the two variants of subAB among 59 eae-negative STEC from large game animals (deer and wild boar) and their meat and meat products in order to assess the role of other species in the epidemiology of subAB-positive, eae-negative STEC. For this approach, the strains were PCR-screened for the presence of subAB, including the specific detection of both allelic variants, for the presence of saa, tia and sab, and for stx subtyping. Overall, subAB genes were detected in 71.2% of the strains: 84.1% of the strains from deer and 33.3% of the strains from wild boar. Most of them (97.6%) possessed subAB2 and most of these subAB2-positive strains (92.7%) were also positive for tia and negative for saa, suggesting the presence of the subAB2-harbouring PAI. Subtype stx2b was present in most of the strains (67.8%) and a statistically significant association could be established between subAB2 and stx2b. Our results suggest that large game animals, mainly deer, may represent an important animal reservoir of subAB2-positive, eae-negative STEC, and also highlight the risk of human infection posed by the consumption of large game meat and meat products. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Occurrence and quantification of Shiga toxin-producing Escherichia coli from food matrices

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    C. Sethulekshmi

    2018-02-01

    Full Text Available Aim: The objective of the study was to detect Shiga toxin-producing Escherichia coli (STEC and develop a quantitative polymerase chain reaction (qPCR assay to quantify the bacterial DNA present in different food matrices. Materials and Methods: A total of 758 samples were collected during a period from January 2015 to December 2016 from Kozhikode, Thrissur, and Alappuzha districts of Kerala. The samples consisted of raw milk (135, pasteurized milk (100, beef (132, buffalo meat (130, chevon (104, beef kheema (115, and beef sausage (42. All the samples collected were subjected to isolation and identification of STEC by conventional culture technique. Confirmation of virulence genes was carried out using PCR. For the quantification of STEC in different food matrices, a qPCR was standardized against stx1 gene of STEC by the construction of standard curve using SYBR green chemistry. Results: The overall occurrence of STEC in raw milk (n=135, beef (n=132, buffalo meat (n=130, chevon (n=104, and beef kheema (n=115 samples collected from Kozhikode, Thrissur, and Alappuzha districts of Kerala was 19.26%, 41.6%, 16.92%, 28.85%, and 41.74%, respectively. PCR revealed the presence of stx1 and stx2 genes in 88.46 and 83.64 and 30.77 and 40.00% of STEC isolates from raw milk and beef samples, respectively, while 100% of the STEC isolates from buffalo beef and beef kheema samples carried stx1 gene. Real-time qPCR assay was used to quantify the bacterial cells present in different food matrices. The standard curve was developed, and the slopes, intercept, and R2 of linear regression curves were -3.10, 34.24, and 0.99, respectively. Conclusion: The considerably high occurrence of STEC in the study confirms the importance of foods of animal origin as a vehicle of infection to humans. In the present study, on comparing the overall occurrence of STEC, the highest percentage of occurrence was reported in beef kheema samples. The study shows the need for rigid food

  20. Effects of zinc supplementation on Shiga toxin 2e-producing Escherichia coli in vitro.

    Science.gov (United States)

    Uemura, Ryoko; Katsuge, Tomoko; Sasaki, Yosuke; Goto, Shinya; Sueyoshi, Masuo

    2017-10-07

    Swine edema disease is caused by Shiga toxin (Stx) 2e-producing Escherichia coli (STEC). Addition of highly concentrated zinc formulations to feed has been used to treat and prevent the disease, but the mechanism of the beneficial effect is unknown. The purpose of the present study was to investigate the effects of highly concentrated zinc formulations on bacterial growth, hemolysin production, and an Stx2e release by STEC in vitro. STEC strain MVH269 isolated from a piglet with edema disease was cultured with zinc oxide (ZnO) or with zinc carbonate (ZnCO 3 ), each at up to 3,000 ppm. There was no effect of zinc addition on bacterial growth. Nonetheless, the cytotoxic activity of Stx2e released into the supernatant was significantly attenuated in the zinc-supplemented media compared to that in the control, with the 50% cytotoxic dose values of 163.2 ± 12.7, 211.6 ± 33.1 and 659.9 ± 84.2 after 24 hr of growth in the presence of ZnO, ZnCO 3 , or no supplemental zinc, respectively. The hemolytic zones around colonies grown on sheep blood agar supplemented with zinc were significantly smaller than those of colonies grown on control agar. Similarly, hemoglobin absorbance after exposure to the supernatants of STEC cultures incubated in sheep blood broth supplemented with zinc was significantly lower than that resulting from exposure to the control supernatant. These in vitro findings indicated that zinc formulations directly impair the factors associated with the virulence of STEC, suggesting a mechanism by which zinc supplementation prevents swine edema disease.

  1. Prevalence and characterization of Shiga Toxin-producing and enteropathogenic Escherichia coli in shellfish-harvesting areas and their watersheds

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    Balière eCharlotte

    2015-12-01

    Full Text Available During a two-year study, the presence of Shiga-toxin producing E. coli (STEC and enteropathogenic E. coli (EPEC was investigated in shellfish (n=238, seawater (n=12 and surface sediment (n=39 collected from three French coastal shellfish-harvesting areas and freshwaters (n=216 in their watersheds. PCR detection of Shiga toxin- (stx1/stx2 and intimin- (eae genes following enrichment from these samples revealed the presence of least one of the stx genes in 30.3% of shellfish batches, 85.9% of freshwater, 41.7% of seawater, and 28.2% of sediment samples, while the eae gene was observed in 74.8%, 100%, 100%, and 43.6% of shellfish batches, freshwater, seawater, and sediment samples, respectively. Twenty-eight STEC and 89 EPEC strains were isolated and analyzed in order to determine their serotype, phylogroup, and genetic relatedness and to evaluate the presence of the saa and ehxA genes encoding the STEC autoagglutinating adhesin and the enterohemolysin A, respectively. Finally, the ability to form biofilms and antimicrobial susceptibility were investigated for a selection of strains. Eighteen serotypes were identified among the STEC isolates and 57 among the EPEC isolates. A high diversity was observed within these strains, as 79 different PFGE patterns and 48 distinguishable sequence types were identified. Strains were found to belong mainly to phylogroups B1 and B2 and virulence was observed to be low as more than 85% of the strains possessed only stx1, stx2 or eae genes. One STEC and several EPEC strains belonged to three of the five highly pathogenic serogroups (i.e., O26, O103, and O145. The subset of strains tested for their capacity to form biofilms was mainly strongly to moderately adherent and more strains formed a strong biofilm at 18°C than at 30°C. Finally, more than 85% of analyzed strains were found to be sensitive to the 16 tested antibiotics. These data suggest the low risk of human infection by STEC if shellfish from these

  2. Evaluation of the performance of the IQ-check kits and the USDA microbiology laboratory guidebook methods for detection of Shiga Toxin-Producing E. coli (STEC) and STEC and Salmonella simultaneously in ground beef

    Science.gov (United States)

    Aims: To evaluate the performance of the IQ-Check kits and the USDA Microbiology Laboratory Guidebook (MLG) methods for detection of the top 7 Shiga toxin-producing E. coli (STEC) (O157:H7, O26, O45, O103, O111, O121, and O145) in ground beef and both STEC and Salmonella in co-inoculated samples. M...

  3. Cellular recovery from exposure to sub-optimal concentrations of AB toxins that inhibit protein synthesis

    Science.gov (United States)

    Shiga toxin 1, exotoxin A, diphtheria toxin and ricin are all AB-type protein toxins that act within the host cytosol to kill the host cell through a pathway involving the inhibition of protein synthesis. It is thought that a single molecule of cytosolic toxin is sufficient to kill the host cell. In...

  4. Expression of Shiga toxin 2e glycosphingolipid receptors of primary porcine brain endothelial cells and toxin-mediated breakdown of the blood-brain barrier.

    Science.gov (United States)

    Meisen, Iris; Rosenbrück, Regina; Galla, Hans-Joachim; Hüwel, Sabine; Kouzel, Ivan U; Mormann, Michael; Karch, Helge; Müthing, Johannes

    2013-06-01

    Shiga toxin (Stx) 2e, released by certain Stx-producing Escherichia coli, is presently the best characterized virulence factor responsible for pig edema disease, which is characterized by hemorrhagic lesions, neurological disorders and often fatal outcomes. Although Stx2e-mediated brain vascular injury is the key event in development of neurologic signs, the glycosphingolipid (GSL) receptors of Stx2e and toxin-mediated impairment of pig brain endothelial cells have not been investigated so far. Here, we report on the detailed structural characterization of Stx2e receptors globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer), which make up the major neutral GSLs in primary porcine brain capillary endothelial cells (PBCECs). Various Gb3Cer and Gb4Cer lipoforms harboring sphingenine (d18:1) or sphinganine (d18:0) and mostly a long-chain fatty acid (C20-C24) were detected. A notable batch-to-batch heterogeneity of primary endothelial cells was observed regarding the extent of ceramide hydroxylation of Gb3Cer or Gb4Cer species. Gb3Cer, Gb4Cer and sphingomyelin preferentially distribute to detergent-resistant membrane fractions and can be considered lipid raft markers in PBCECs. Moreover, we employed an in vitro model of the blood-brain barrier (BBB), which exhibited strong cytotoxic effects of Stx2e on the endothelial monolayer and a rapid collapse of the BBB. These data strongly suggest the involvement of Stx2e in cerebral vascular damage with resultant neurological disturbance characteristic of edema disease.

  5. Characteristics of Clinical Shiga Toxin-Producing Escherichia coli Isolated from British Columbia

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    Kevin J. Allen

    2013-01-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC are significant public health threats. Although STEC O157 are recognized foodborne pathogens, non-O157 STEC are also important causes of human disease. We characterized 10 O157:H7 and 15 non-O157 clinical STEC derived from British Columbia (BC. Eae, hlyA, and stx were more frequently observed in STEC O157, and 80 and 100% of isolates possessed stx1 and stx2, respectively. In contrast, stx1 and stx2 occurred in 80 and 40% of non-O157 STEC, respectively. Comparative genomic fingerprinting (CGF revealed three distinct clusters (C. STEC O157 was identified as lineage I (LI; LSPA-6 111111 and clustered as a single group (C1. The cdi gene previously observed only in LII was seen in two LI O157 isolates. CGF C2 strains consisted of diverse non-O157 STEC while C3 included only O103:H25, O118, and O165 serogroup isolates. With the exception of O121 and O165 isolates which were similar in virulence gene complement to STEC O157, C1 O157 STEC produced more Stx2 than non-O157 STEC. Antimicrobial resistance (AMR screening revealed resistance or reduced sensitivity in all strains, with higher levels occurring in non-O157 STEC. One STEC O157 isolate possessed a mobile blaCMY-2 gene transferrable across genre via conjugation.

  6. Detection, Characterization, and Typing of Shiga Toxin-Producing Escherichia coli.

    Science.gov (United States)

    Parsons, Brendon D; Zelyas, Nathan; Berenger, Byron M; Chui, Linda

    2016-01-01

    Shiga toxin-producing Escherichia coli (STEC) are responsible for gastrointestinal diseases reported in numerous outbreaks around the world. Given the public health importance of STEC, effective detection, characterization and typing is critical to any medical laboratory system. While non-O157 serotypes account for the majority of STEC infections, frontline microbiology laboratories may only screen for STEC using O157-specific agar-based methods. As a result, non-O157 STEC infections are significantly under-reported. This review discusses recent advances on the detection, characterization and typing of STEC with emphasis on work performed at the Alberta Provincial Laboratory for Public Health (ProvLab). Candidates for the detection of all STEC serotypes include chromogenic agars, enzyme immunoassays (EIA) and quantitative real time polymerase chain reaction (qPCR). Culture methods allow further characterization of isolates, whereas qPCR provides the greatest sensitivity and specificity, followed by EIA. The virulence gene profiles using PCR arrays and stx gene subtypes can subsequently be determined. Different non-O157 serotypes exhibit markedly different virulence gene profiles and a greater prevalence of stx1 than stx2 subtypes compared to O157:H7 isolates. Finally, recent innovations in whole genome sequencing (WGS) have allowed it to emerge as a candidate for the characterization and typing of STEC in diagnostic surveillance isolates. Methods of whole genome analysis such as single nucleotide polymorphisms and k-mer analysis are concordant with epidemiological data and standard typing methods, such as pulsed-field gel electrophoresis and multiple-locus variable number tandem repeat analysis while offering additional strain differentiation. Together these findings highlight improved strategies for STEC detection using currently available systems and the development of novel approaches for future surveillance.

  7. Detection, Characterization and Typing of Shiga toxin-producing Escherichia coli.

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    Brendon David Parsons

    2016-04-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC are responsible for gastrointestinal diseases reported in numerous outbreaks around the world. Given the public health importance of STEC, effective detection, characterization and typing is critical to any medical laboratory system. While non-O157 serotypes account for the majority of STEC infections, frontline microbiology laboratories may only screen for STEC using O157-specific agar-based methods. As a result, non-O157 STEC infections are significantly under-reported. This review discusses recent advances on the detection, characterization and typing of STEC with emphasis on work performed at the Alberta Provincial Laboratory for Public Health (ProvLab. Candidates for the detection of all STEC serotypes include chromogenic agars, enzyme immunoassays (EIA and real-time polymerase chain reaction (qPCR. Culture methods allow further characterization of isolates, whereas qPCR provides the greatest sensitivity and specificity, followed by EIA. The virulence gene profiles using PCR arrays and stx gene subtypes can subsequently be determined. Different non-O157 serotypes exhibit markedly different virulence gene profiles and a greater prevalence of stx1 than stx2 subtypes compared to O157:H7 isolates. Finally, recent innovations in whole genome sequencing (WGS have allowed it to emerge as a candidate for the characterization and typing of STEC in diagnostic surveillance isolates. Methods of whole genome analysis such as single nucleotide polymorphisms and k-mer analysis are concordant with epidemiological data and standard typing methods, such as pulsed-field gel electrophoresis and multiple-locus variable number tandem repeat analysis while offering additional strain differentiation. Together these findings highlight improved strategies for STEC detection using currently available systems and the development of novel approaches for future surveillance.

  8. Contribution of Urease to Colonization by Shiga Toxin-Producing Escherichia coli

    Science.gov (United States)

    Steyert, Susan R.

    2012-01-01

    Shiga toxin-producing Escherichia coli (STEC) is a food-borne pathogen with a low infectious dose that colonizes the colon in humans and can cause severe clinical manifestations such as hemolytic-uremic syndrome. The urease enzyme, encoded in the STEC chromosome, has been demonstrated to act as a virulence factor in other bacterial pathogens. The NH3 produced as urease hydrolyzes urea can aid in buffering bacteria in acidic environments as well as provide an easily assimilated source of nitrogen that bacteria can use to gain a metabolic advantage over intact microflora. Here, we explore the role of urease in STEC pathogenicity. The STEC urease enzyme exhibited maximum activity near neutral pH and during the stationary-growth phase. Experiments altering growth conditions performed with three phylogenetically distinct urease-positive strains demonstrated that the STEC ure gene cluster is inducible by neither urea nor pH but does respond to nitrogen availability. Quantitative reverse transcription-PCR (qRT-PCR) data indicate that nitrogen inhibits the transcriptional response. The deletion of the ure gene locus was constructed in STEC strain 88-0643, and the ure mutant was used with the wild-type strain in competition experiments in mouse models to examine the contribution of urease. The wild-type strain was twice as likely to survive passage through the acidic stomach and demonstrated an enhanced ability to colonize the intestinal tract compared to the ure mutant strain. These in vivo experiments reveal that, although the benefit STEC gains from urease expression is modest and not absolutely required for colonization, urease can contribute to the pathogenicity of STEC. PMID:22665380

  9. Agitation down-regulates immunoglobulin binding protein EibG expression in Shiga toxin-producing Escherichia coli (STEC.

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    Thorsten Kuczius

    Full Text Available Shiga toxin (Stx-producing Escherichia coli (STEC carrying eibG synthesize Escherichia coli immunoglobulin binding protein (EibG. EibG nonspecifically binds to immunoglobulins and tends to aggregate in multimers but is poorly expressed in wild-type strains. To study synthesis of the proteins and their regulation in the pathogens, we identified natural growth conditions that increased EibG synthesis. EibG proteins as well as corresponding mRNA were highly expressed under static growth conditions while shearing stress created by agitation during growth repressed protein synthesis. Further regulation effects were driven by reduced oxygen tension, and pH up-regulated EibG expression, but to a lesser extent than growth conditions while decreased temperature down-regulated EibG. Bacteria with increased EibG expression during static growth conditions showed a distinct phenotype with chain formation and biofilm generation, which disappeared with motion. High and low EibG expression was reversible indicating a process with up- and down-regulation of the protein expression. Our findings indicate that shear stress represses EibG expression and might reduce bacterial attachments to cells and surfaces.

  10. Effect of carvacrol on O157 and non-O157 Shiga toxin producing Escherichia coli

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    Alexandros Stratakos

    2017-06-01

    Full Text Available Shiga toxin Escherichia coli (STEC strains are important foodborne bacteria linked to diarrhea, enteritis, hemolytic-uremic syndrome and in some cases death. E. coli O157:H7 is the most common strain amongst STECs however non-O157 STECs have been connected with several outbreaks on an international level.  The use of natural plant extracts to reduce the risk from foodborne pathogens is gaining increasing importance. Therefore in this study, we tested the antibacterial effect of carvacrol, a major component of oregano essential oil, on E. coli serogroups O157, O26, O45, O103, O111, O121, O145 as well as serogroup O104 responsible for the massive outbreak in Germany in 2011. Carvacrol showed antibacterial effect on all strains tested. The relative electric conductivity was assessed in order to investigate the changes in membrane permeability and thus to investigate the antimicrobial mechanism of carvacrol. Results showed that the relative conductivity increased with increasing concentrations of carvacrol which showed that there was an increasing leakage of electrolytes due to disruption of the cell membrane. The data presented here revealed that carvacrol has the potential to be used as a natural antimicrobial against STECs.

  11. Improved traceability of Shiga-toxin-producing Escherichia coli using CRISPRs for detection and typing.

    Science.gov (United States)

    Delannoy, Sabine; Beutin, Lothar; Fach, Patrick

    2016-05-01

    Among strains of Shiga-toxin-producing Escherichia coli (STEC), seven serogroups (O26, O45, O103, O111, O121, O145, and O157) are frequently associated with severe clinical illness in humans. The development of methods for their reliable detection from complex samples such as food has been challenging thus far, and is currently based on the PCR detection of the major virulence genes stx1, stx2, and eae, and O-serogroup-specific genes. However, this approach lacks resolution. Moreover, new STEC serotypes are continuously emerging worldwide. For example, in May 2011, strains belonging to the hitherto rarely detected STEC serotype O104:H4 were identified as causative agents of one of the world's largest outbreak of disease with a high incidence of hemorrhagic colitis and hemolytic uremic syndrome in the infected patients. Discriminant typing of pathogens is crucial for epidemiological surveillance and investigations of outbreaks, and especially for tracking and tracing in case of accidental and deliberate contamination of food and water samples. Clustered regularly interspaced short palindromic repeats (CRISPRs) are composed of short, highly conserved DNA repeats separated by unique sequences of similar length. This distinctive sequence signature of CRISPRs can be used for strain typing in several bacterial species including STEC. This review discusses how CRISPRs have recently been used for STEC identification and typing.

  12. Influence of yogurt fermentation and refrigerated storage on the stability of protein toxin contaminants.

    Science.gov (United States)

    Jackson, Lauren S; Triplett, Odbert A; Tolleson, William H

    2015-06-01

    Dairy products sold in a ready-to-eat form present the risk that adulterants persisting through manufacturing, storage, and distribution would reach consumers. Pathogenic microbes, including shigatoxigenic strains of Escherichia coli and the toxins they produce, are common food safety hazards associated with dairy products. Ricin and abrin are plant-derived ribosome-inactivating protein toxins related to the shiga-like toxins produced by E. coli. Limited information exists on the effects of manufacturing processes on the stabilities of these heat-resistant ribosome-inactivating proteins in the presence of foods. The goal of this study was to determine how typical yogurt manufacturing and storage processes influence ribosome-inactivating protein toxins. Ricin and abrin were added to skim or whole milk and batch pasteurized. Complete inactivation of both toxins was observed after 30 minutes at 85 °C. If the toxins were added after pasteurization, the levels of ricin and abrin in yogurt and their cytotoxic activities did not change significantly during fermentation or refrigerated storage for 4 weeks. The activities of ricin and abrin were inhibited by skim milk, nonfat yogurt, whole milk, and whole milk yogurt. The results showed minimal effects of the toxins on yogurt pH and %titratable acidity but inhibitory effects of yogurt on toxin activity. Published by Elsevier Ltd.

  13. Monoclonal antibodies and toxins--a perspective on function and isotype.

    Science.gov (United States)

    Chow, Siu-Kei; Casadevall, Arturo

    2012-06-01

    Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins--Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)--and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions.

  14. Characteristics of Shiga toxin-producing Escherichia coli from meat and milk products of different origins and association with food producing animals as main contamination sources.

    Science.gov (United States)

    Martin, Annett; Beutin, Lothar

    2011-03-15

    Shiga toxin-producing strains of Escherichia coli (STEC) cause diarrhoea and haemorrhagic colitis in humans. Most human infections are attributed to consumption of STEC contaminated foodstuff. Food producing animals constitute important reservoirs of STEC and serve as source of food contamination. In this study, we have analyzed 593 foodborne STEC strains for their serotypes and for nine virulence genes (stx1, stx1c, stx1d, stx2, stx2b, stx2e, stx2g, E-hly and eae). The 593 STEC strains grouped into 215 serotypes, and 123 serotypes (57.2%) were represented each by only one STEC isolate. Fifteen serotypes (7.0%) were attributed to 198 (33.3%) of the 593 STEC strains. The foodborne STEC were grouped into different categories in relation to the species of the food producing animal (cattle, pigs, sheep, goats, red deer, wild-boar and hare). Univariate and multivariate statistical analyses revealed significant similarities between the animal origin of the food and the virulence markers of foodborne STEC. Significant associations (pfood producing animals. Virulence profiles and serotypes of STEC from food showed remarkable similarities to those of faecal STEC that were from the same animal species. The findings from our study clearly indicate that the food producing animals represent the most important source for the entry of STEC in the food chain. Sound hygiene measures implemented at critical stages of food production (milking, slaughtering, and evisceration) should be most effective in reducing the frequency of STEC contamination of food derived from domestic and wildlife animals. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. VIABILIDADE DE Escherichia coli PRODUTORA DE TOXINA SHIGA (STEC NÃO-O157 EM QUEIJO TIPO MINAS FRESCAL. VIABILITY OF NON-O157 SHIGA TOXIN-PRODUCING Escherichia coli (STEC IN MINAS FRESCAL CHEESE

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    Tammy Priscila Chioda

    2009-07-01

    Full Text Available Escherichia coli, produtora de toxina Shiga (STEC, um patógeno emergente capaz de causar diarreia, colite hemorrágica e síndrome hemolítica urêmica em humanos, representa um grave problema de saúde pública em todo o mundo. O principal reservatório de STEC são os bovinos. STEC são transmitidas aos humanos, principalmente através de alimentos contaminados, destacando-se aqueles de origem bovina como carne, leite e seus derivados. O objetivo deste trabalho foi avaliar a viabilidade de STEC não-O157 em queijo minas frescal preparado com leite artificialmente contaminado com diferentes cepas dessas bactérias. Os queijos foram mantidos a 4°C e analisados no 1º, 2º, 4º, 6º e 10º dias de estocagem. As cepas de STEC mantiveram-se viáveis em 100% (32/32 dos queijos mantidos sob refrigeração por até dez dias. Os resultados mostram que o queijo minas pode ser veículo de transmissão de STEC. Recomenda-se a adoção de métodos higiênicos e sanitários desde a ordenha até o processo de produção do queijo para reduzir a possibilidade de contaminação com STEC.

    PALAVRAS-CHAVES: PCR, queijo minas, segurança alimentar, STEC.

    Shiga toxin-producing Escherichia coli (STEC an emergent foodborne pathogen that cause diarrhea, hemorrhagic colitis and haemolytic uremic syndrome in humans, represents a public health problem all over the world. Cattle are the main source of STEC. STEC are transmitted to humans by contaminated food, mainly those of bovine origin as meat and dairy products. This study aimed evaluates the non-O157 STEC viability of artificially inoculated in the milk used for the Minas Frescal cheese’s production. The cheese was kept at 4°C and analyzed at 1st, 2nd, 4th, 6th and 10th days after its production. 100% (32/32 of the cheese storad under refrigeration during 10 days had been the STEC strains viable. These results show that minas frescal cheese can transmit STEC. The adoption of good

  16. Taxonomy Meets Public Health: The Case of Shiga Toxin-Producing Escherichia coli.

    Science.gov (United States)

    Scheutz, Flemming

    2014-06-01

    To help assess the clinical and public health risks associated with different Shiga toxin-producing Escherichia coli (STEC) strains, an empirical classification scheme was used to classify STEC into five "seropathotypes" (seropathotype A [high risk] to seropathotypes D and E [minimal risk]). This definition is of considerable value in cases of human infection but is also problematic because not all STEC infections are fully characterized and coupled to reliable clinical information. Outbreaks with emerging hybrid strains continuously challenge our understanding of virulence potential and may result in incorrect classification of specific pathotypes; an example is the hybrid strain that caused the 2011 outbreak in Germany, STEC/EAggEC O104:H4, which may deserve an alternative seropathotype designation. The integration of mobile virulence factors in the stepwise and parallel evolution of pathogenic lineages of STEC collides with the requirements of a good taxonomy, which separates elements of each group into subgroups that are mutually exclusive, unambiguous, and, together, include all possibilities. The concept of (sero)-pathotypes is therefore challenged, and the need to identify factors of STEC that absolutely predict the potential to cause human disease is obvious. Because the definition of hemolytic-uremic syndrome (HUS) is distinct, a basic and primary definition of HUS-associated E. coli (HUSEC) for first-line public health action is proposed: stx2 in a background of an eae- or aggR-positive E. coli followed by a second-line subtyping of stx genes that refines the definition of HUSEC to include only stx2a and stx2d. All other STEC strains are considered "low-risk" STEC.

  17. Genetic characterization of Shiga toxin-producing Escherichia coli O26:H11 strains isolated from animal, food, and clinical samples

    Science.gov (United States)

    Krüger, Alejandra; Lucchesi, Paula M. A.; Sanso, A. Mariel; Etcheverría, Analía I.; Bustamante, Ana V.; Burgán, Julia; Fernández, Luciana; Fernández, Daniel; Leotta, Gerardo; Friedrich, Alexander W.; Padola, Nora L.; Rossen, John W. A.

    2015-01-01

    The Shiga-toxin producing Escherichia coli (STEC) may cause serious illness in human. Here we analyze O26:H11 strains known to be among the most reported STEC strains causing human infections. Genetic characterization of strains isolated from animal, food, and clinical specimens in Argentina showed that most carried either stx1a or stx2a subtypes. Interestingly, stx2a-positive O26:H11 rarely isolated from cattle in other countries showed to be an important proportion of O26:H11 strains circulating in cattle and food in our region. Seventeen percent of the isolates harbored more than one gene associated with antimicrobial resistance. In addition to stx, all strains contained the virulence genes eae-β, tir, efa, iha, espB, cif, espA, espF, espJ, nleA, nleB, nleC, and iss; and all except one contained ehxA, espP, and cba genes. On the other hand, toxB and espI genes were exclusively observed in stx2-positive isolates, whereas katP was only found in stx1a-positive isolates. Our results show that O26:H11 STEC strains circulating in Argentina, including those isolated from humans, cattle, and meat products, present a high pathogenic potential, and evidence that cattle can be a reservoir of O26:H11 strains harboring stx2a. PMID:26539413

  18. DNA fingerprinting of Shiga-toxin producing Escherichia coli O157 based on Multiple-Locus Variable-Number Tandem-Repeats Analysis (MLVA

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    Vardund Traute

    2003-12-01

    Full Text Available Abstract Background The ability to react early to possible outbreaks of Escherichia coli O157:H7 and to trace possible sources relies on the availability of highly discriminatory and reliable techniques. The development of methods that are fast and has the potential for complete automation is needed for this important pathogen. Methods In all 73 isolates of shiga-toxin producing E. coli O157 (STEC were used in this study. The two available fully sequenced STEC genomes were scanned for tandem repeated stretches of DNA, which were evaluated as polymorphic markers for isolate identification. Results The 73 E. coli isolates displayed 47 distinct patterns and the MLVA assay was capable of high discrimination between the E. coli O157 strains. The assay was fast and all the steps can be automated. Conclusion The findings demonstrate a novel high discriminatory molecular typing method for the important pathogen E. coli O157 that is fast, robust and offers many advantages compared to current methods.

  19. Interplay between toxin transport and flotillin localization

    DEFF Research Database (Denmark)

    Pust, Sascha; Dyve, Anne Berit; Torgersen, Maria L

    2010-01-01

    The flotillin proteins are localized in lipid domains at the plasma membrane as well as in intracellular compartments. In the present study, we examined the importance of flotillin-1 and flotillin-2 for the uptake and transport of the bacterial Shiga toxin (Stx) and the plant toxin ricin and we...... for flotillin-1 or -2. However, the Golgi-dependent sulfation of both toxins was significantly reduced in flotillin knockdown cells. Interestingly, when the transport of ricin to the ER was investigated, we obtained an increased mannosylation of ricin in flotillin-1 and flotillin-2 knockdown cells. The toxicity...... of both toxins was twofold increased in flotillin-depleted cells. Since BFA (Brefeldin A) inhibits the toxicity even in flotillin knockdown cells, the retrograde toxin transport is apparently still Golgi-dependent. Thus, flotillin proteins regulate and facilitate the retrograde transport of Stx and ricin....

  20. Diversity of Shiga Toxin-Producing Escherichia coli (STEC) O26:H11 Strains Examined via stx Subtypes and Insertion Sites of Stx and EspK Bacteriophages

    Science.gov (United States)

    Bonanno, Ludivine; Loukiadis, Estelle; Mariani-Kurkdjian, Patricia; Oswald, Eric; Garnier, Lucille; Michel, Valérie

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) is a food-borne pathogen that may be responsible for severe human infections. Only a limited number of serotypes, including O26:H11, are involved in the majority of serious cases and outbreaks. The main virulence factors, Shiga toxins (Stx), are encoded by bacteriophages. Seventy-four STEC O26:H11 strains of various origins (including human, dairy, and cattle) were characterized for their stx subtypes and Stx phage chromosomal insertion sites. The majority of food and cattle strains possessed the stx1a subtype, while human strains carried mainly stx1a or stx2a. The wrbA and yehV genes were the main Stx phage insertion sites in STEC O26:H11, followed distantly by yecE and sbcB. Interestingly, the occurrence of Stx phages inserted in the yecE gene was low in dairy strains. In most of the 29 stx-negative E. coli O26:H11 strains also studied here, these bacterial insertion sites were vacant. Multilocus sequence typing of 20 stx-positive or stx-negative E. coli O26:H11 strains showed that they were distributed into two phylogenetic groups defined by sequence type 21 (ST21) and ST29. Finally, an EspK-carrying phage was found inserted in the ssrA gene in the majority of the STEC O26:H11 strains but in only a minority of the stx-negative E. coli O26:H11 strains. The differences in the stx subtypes and Stx phage insertion sites observed in STEC O26:H11 according to their origin might reflect that strains circulating in cattle and foods are clonally distinct from those isolated from human patients. PMID:25819955

  1. Effects of subinhibitory concentrations of antimicrobial agents on Escherichia coli O157:H7 Shiga toxin release and role of the SOS response.

    Science.gov (United States)

    Nassar, Farah J; Rahal, Elias A; Sabra, Ahmad; Matar, Ghassan M

    2013-09-01

    Treatment of Escherichia coli O157:H7 by certain antimicrobial agents often exacerbates the patient's condition by increasing either the release of preformed Shiga toxins (Stx) upon cell lysis or their production through the SOS response-triggered induction of Stx-producing prophages. Recommended subinhibitory concentrations (sub-MICs) of azithromycin (AZI), gentamicin (GEN), imipenem (IMI), and rifampicin (RIF) were evaluated in comparison to norfloxacin (NOR), an SOS-inducer, to assess the role of the SOS response in Stx release. Relative expression of recA (SOS-inducer), Q (late antitermination gene of Stx-producing prophage), stx1, and stx2 genes was assessed at two sub-MICs of the antimicrobials for two different strains of E. coli O157:H7 using reverse transcription-real-time polymerase chain reaction. Both strains at the two sub-MICs were also subjected to Western blotting for LexA protein expression and to reverse passive latex agglutination for Stx detection. For both strains at both sub-MICs, NOR and AZI caused SOS-induced Stx production (high recA, Q, and stx2 gene expression and high Stx2 production), so they should be avoided in E. coli O157:H7 treatment; however, sub-MICs of RIF and IMI induced Stx2 production in an SOS-independent manner except for one strain at the first twofold dilution below MIC of RIF where Stx2 production decreased. Moreover, GEN caused somewhat increased Stx2 production due to its mode of action rather than any effect on gene expression. The choice of antimicrobial therapy should rely on the antimicrobial mode of action, its concentration, and on the nature of the strain.

  2. Comparison of the fate of the top six non-O157 shiga-toxin producing Escherichia coli (STEC) and E. coli O157:H7 during the manufacture of dry fermented sausages.

    Science.gov (United States)

    Balamurugan, S; Ahmed, Rafath; Gao, Anli; Strange, Phil

    2017-10-16

    The study examined the relative fate of the top six non-O157 shiga-toxin producing Escherichia coli (STEC) and E. coli O157:H7 during the manufacture of dry fermented sausages (DFS). Three separate batches of sausages containing a five-strain cocktail for each serogroup and uninoculated control were manufactured and subjected to identical fermentation, maturation and dry curing conditions. Changes in physicochemical properties and inoculated STEC numbers were enumerated during the DFS production stages and log reduction and log reduction rates were calculated. Inoculation of very high concentrations (8logCFUg -1 ) of STEC in the sausage batter did not significantly (P>0.05) affect the changes in the pH, a w , moisture, protein, fat content compared to the uninoculated DFS. There was a significant (P0.05) from each other. These results indicate that the lethality of DFS production processes observed against E. coli O157:H7 would result in a similar inactivation of the top six non-O157 STEC. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

  3. Prevalence of shiga toxin producing Escherichia coli, Salmonella enterica, and Listeria monocytogenes at public access watershed sites in a California Central Coast agricultural region.

    Science.gov (United States)

    Cooley, Michael B; Quiñones, Beatriz; Oryang, David; Mandrell, Robert E; Gorski, Lisa

    2014-01-01

    Produce contaminated with enteric pathogens is a major source of foodborne illness in the United States. Lakes, streams, rivers, and ponds were sampled with Moore swabs bi-monthly for over 2 years at 30 locations in the vicinity of a leafy green growing region on the Central California Coast and screened for Shiga toxin producing Escherichia coli (STEC), Salmonella enterica, and Listeria monocytogenes to evaluate the prevalence and persistence of pathogen subtypes. The prevalence of STEC from 1386 samples was 11%; 110 samples (8%) contained E. coli O157:H7 with the highest prevalence occurring close to cattle operations. Non-O157 STEC isolates represented major clinical O-types and 57% contained both shiga toxin types 1 and 2 and intimin. Multiple Locus Variable Number Tandem Repeat Analysis of STEC isolates indicated prevalent strains during the period of study. Notably, Salmonella was present at high levels throughout the sampling region with 65% prevalence in 1405 samples resulting in 996 isolates with slightly lower prevalence in late autumn. There were 2, 8, and 14 sites that were Salmonella-positive over 90, 80, and 70% of the time, respectively. The serotypes identified most often were 6,8:d:-, Typhimurium, and Give. Interestingly, analysis by Pulsed Field Gel Electrophoresis indicated persistence and transport of pulsotypes in the region over several years. In this original study of L. monocytogenes in the region prevalence was 43% of 1405 samples resulting in 635 individual isolates. Over 85% of the isolates belonged to serotype 4b with serotypes 1/2a, 1/2b, 3a, 4d with 4e representing the rest, and there were 12 and 2 sites that were positive over 50 and 80% of the time, respectively. Although surface water is not directly used for irrigation in this region, transport to the produce can occur by other means. This environmental survey assesses initial contamination levels toward an understanding of transport leading to produce recalls or outbreaks.

  4. Conditional Function of Autoaggregative Protein Cah and Common cah Mutations in Shiga Toxin-Producing Escherichia coli.

    Science.gov (United States)

    Carter, Michelle Qiu; Brandl, Maria T; Kudva, Indira T; Katani, Robab; Moreau, Matthew R; Kapur, Vivek

    2018-01-01

    Cah is a calcium-binding autotransporter protein involved in autoaggregation and biofilm formation. Although cah is widespread in Shiga toxin-producing Escherichia coli (STEC), we detected mutations in cah at a frequency of 31.3% in this pathogen. In STEC O157:H7 supershedder strain SS17, a large deletion results in a smaller coding sequence, encoding a protein lacking the C-terminal 71 amino acids compared with Cah in STEC O157:H7 strain EDL933. We examined the function of Cah in biofilm formation and host colonization to better understand the selective pressures for cah mutations. EDL933-Cah played a conditional role in biofilm formation in vitro : it enhanced E. coli DH5α biofilm formation on glass surfaces under agitated culture conditions that prevented autoaggregation but inhibited biofilm formation under hydrostatic conditions that facilitated autoaggregation. This function appeared to be strain dependent since Cah-mediated biofilm formation was diminished when an EDL933 cah gene was expressed in SS17. Deletion of cah in EDL933 enhanced bacterial attachment to spinach leaves and altered the adherence pattern of EDL933 to bovine recto-anal junction squamous epithelial (RSE) cells. In contrast, in trans expression of EDL933 cah in SS17 increased its attachment to leaf surfaces, and in DH5α, it enhanced its adherence to RSE cells. Hence, the ecological function of Cah appears to be modulated by environmental conditions and other bacterial strain-specific properties. Considering the prevalence of cah in STEC and its role in attachment and biofilm formation, cah mutations might be selected in ecological niches in which inactivation of Cah would result in an increased fitness in STEC during colonization of plants or animal hosts. IMPORTANCE Shiga toxin-producing Escherichia coli (STEC) harbors genes encoding diverse adhesins, and many of these are known to play an important role in bacterial attachment and host colonization. We demonstrated here that the

  5. Lysogeny with Shiga Toxin 2-Encoding Bacteriophages Represses Type III Secretion in Enterohemorrhagic Escherichia coli

    Science.gov (United States)

    Xu, Xuefang; McAteer, Sean P.; Tree, Jai J.; Shaw, Darren J.; Wolfson, Eliza B. K.; Beatson, Scott A.; Roe, Andrew J.; Allison, Lesley J.; Chase-Topping, Margo E.; Mahajan, Arvind; Tozzoli, Rosangela; Woolhouse, Mark E. J.; Morabito, Stefano; Gally, David L.

    2012-01-01

    Lytic or lysogenic infections by bacteriophages drive the evolution of enteric bacteria. Enterohemorrhagic Escherichia coli (EHEC) have recently emerged as a significant zoonotic infection of humans with the main serotypes carried by ruminants. Typical EHEC strains are defined by the expression of a type III secretion (T3S) system, the production of Shiga toxins (Stx) and association with specific clinical symptoms. The genes for Stx are present on lambdoid bacteriophages integrated into the E. coli genome. Phage type (PT) 21/28 is the most prevalent strain type linked with human EHEC infections in the United Kingdom and is more likely to be associated with cattle shedding high levels of the organism than PT32 strains. In this study we have demonstrated that the majority (90%) of PT 21/28 strains contain both Stx2 and Stx2c phages, irrespective of source. This is in contrast to PT 32 strains for which only a minority of strains contain both Stx2 and 2c phages (28%). PT21/28 strains had a lower median level of T3S compared to PT32 strains and so the relationship between Stx phage lysogeny and T3S was investigated. Deletion of Stx2 phages from EHEC strains increased the level of T3S whereas lysogeny decreased T3S. This regulation was confirmed in an E. coli K12 background transduced with a marked Stx2 phage followed by measurement of a T3S reporter controlled by induced levels of the LEE-encoded regulator (Ler). The presence of an integrated Stx2 phage was shown to repress Ler induction of LEE1 and this regulation involved the CII phage regulator. This repression could be relieved by ectopic expression of a cognate CI regulator. A model is proposed in which Stx2-encoding bacteriophages regulate T3S to co-ordinate epithelial cell colonisation that is promoted by Stx and secreted effector proteins. PMID:22615557

  6. Membrane invagination induced by Shiga toxin B-subunit

    DEFF Research Database (Denmark)

    Pezeshkian, W.; Hansen, Allan Grønhøj; Johannes, Ludger

    2016-01-01

    -atom molecular dynamics and Monte Carlo simulations we show that the molecular architecture of STxB enables the following sequence of events: the Gb3 binding sites on STxB are arranged such that tight avidity-based binding results in a small increment of local curvature. Membrane-mediated clustering of several...... toxin molecules then creates a tubular membrane invagination that drives toxin entry into the cell. This mechanism requires: (1) a precise molecular architecture of the STxB binding sites; (2) a fluid bilayer in order for the tubular invagination to form. Although, STxB binding to the membrane requires...... specific interactions with Gb3 lipids, our study points to a generic molecular design principle for clathrin-independent endocytosis of nanoparticles....

  7. Quantification of plasmid DNA reference materials for Shiga toxin-producing Escherichia coli based on UV, HR-ICP-MS and digital PCR.

    Science.gov (United States)

    Liang, Wen; Xu, Li; Sui, Zhiwei; Li, Yan; Li, Lanying; Wen, Yanli; Li, Chunhua; Ren, Shuzhen; Liu, Gang

    2016-01-01

    The accuracy and metrology traceability of DNA quantification is becoming a critical theme in many fields, including diagnosis, forensic analysis, microorganism detection etc. Thus the research of DNA reference materials (RMs) and consistency of DNA quantification methods has attracted considerable research interest. In this work, we developed 3 plasmid candidate RMs, containing 3 target genes of Escherichia coli O157:H7 (E. coli O157:H7) and other Shiga toxin-producing Escherichia coli (STEC): stx1, stx2, and fliC (h7) respectively. Comprehensive investigation of the plasmid RMs was performed for their sequence, purity, homogeneity and stability, and then the concentration was quantified by three different methods: ultraviolet spectrophotometer (UV), high resolution inductively coupled plasma mass spectrometry (HR-ICP-MS) and digital PCR. As a routinely applied method for DNA analysis, UV was utilized for the quantification (OD260) and purity analysis for the plasmids. HR-ICP-MS quantified the plasmid DNA through analysing the phosphorus in DNA molecules. Digital PCR distributed the DNA samples onto a microarray chip containing thousands of reaction chambers, and quantified the DNA copy numbers by analysing the number of positive signals without any calibration curves needed. Based on the high purification of the DNA reference materials and the optimization of dPCR analysis, we successfully achieved good consistency between UV, HR-ICP-MS and dPCR, with relative deviations lower than 10 %. We then performed the co-quantification of 3 DNA RMs with three different methods together, and the uncertainties of their concentration were evaluated. Finally, the certified values and expanded uncertainties for 3 DNA RMs (pFliC, pStx1 and pStx2) were (1.60 ± 0.10) × 10(10) copies/μL, (1.53 ± 0.10) × 10(10) copies/μL and (1.70 ± 0.11) × 10(10) copies/μL respectively.Graphical abstractWe developed 3 plasmid candidate RMs, containing 3 target genes of

  8. Survey of Shiga toxin-producing Escherichia coli O157:H7 in urban pigeons (Columba livia in the city of Napoli, Italy

    Directory of Open Access Journals (Sweden)

    Alessandro Fioretti

    2010-01-01

    Full Text Available Recently, several studies have demonstrated that pigeon is an important reservoir of Shiga toxin-producing E. coli O157:H7. The aim of this study was to evaluate the presence of this pathogen in urban pigeons in the city of Napoli. The sampling was carried out during the period November 2005/July 2006. The city was subdivided in 56 quadrants by Geographical Information System. Each quadrant was analysed three times. From each quadrant, 3 pigeons were analysed by cloacal swabs. A total of 504 cloacal swabs was obtained. We isolated four E. coli O157:H7 strains. By multiplex PCR, all strains carried eae and stx2 genes, whereas only one strain carried the stx1 gene. 2/4 isolated strains carried hly gene which is considered a hallmark of human pathogenic strains. Our results indicate that pigeon faces are a source of E. coli O157:H7 for birds, mammals and humans.

  9. Dynamics of plc gene transcription and α-toxin production during growth of Clostridium perfringens strains with contrasting α-toxin production

    DEFF Research Database (Denmark)

    Abildgaard, Lone; Schramm, Andreas; Rudi, Knut

    2009-01-01

    The aim of the present study was to investigate transcription dynamics of the α-toxin-encoding plc gene relative to two housekeeping genes (gyrA and rplL) in batch cultures of three Clostridium perfringens strains with low, intermediate, and high levels of α-toxin production, respectively. The plc...... transcript level was always low in the low α-toxin producing strain. For the two other strains, plc transcription showed an inducible pattern and reached a maximum level in the late exponential growth phase. The transcription levels were however inversely correlated to α-toxin production for the two strains....... We propose that this discrepancy is due to differences in plc translation rates between the strains and that strain-specific translational rates therefore must be determined before α-toxin production can be extrapolated from transcript levels in C. perfringens....

  10. Prevalence, quantification and isolation of pathogenic shiga toxin Escherichia coli O157:H7 along the production and supply chain of pork around Hubei Province of China.

    Science.gov (United States)

    Khan, Sher Bahadar; Zou, Geng; Xiao, Ran; Cheng, Yuting; Rehman, Zia Ur; Ali, Sher; Memon, Atta Muhammad; Fahad, Shah; Ahmad, Irshad; Zhou, Rui

    2018-02-01

    Shiga toxin Escherichia coli (STEC) O157:H7 is an important zoonotic food borne pathogen causing gastroenteritis that may lead to life threatening hemorragic colitis (HC) and hemorrhagic uremic syndrome (HUS). 325 meat and tissue samples were tested for enumeration of O157:H7 strains using most probable number (MPN)-PCR targeting their specific genes flicH7 and rfbO157 followed by isolation, sereotyping and pathogenicity testing. The overall prevalence of O157:H7 was 41.3% (134/325) along the production and supply chain of pork (PSCP), being higher in supply chain (59%, 118/200) as compared to pig farms (12.8%, 16/125). Along the PSCP, the highest prevalence was found in slaughter houses (86.25%, 69/80) followed by wet- (53.3%, 32/60) and super-markets (28.3%, 17/60). The MPN values ranged from 3 to 1100 MPN/g in overall positive samples, being higher in slaughter houses followed by wet and super markets. Except from intestine and meat samples of slaughter house, the MPN was found higher in summer as compared to winter samples. Eight STEC O157:H7 isolated from meat and liver samples were tested in Balb/C mice for pathogenicity. After development of clinical signs and symptoms, 50-83.3% mortality was produced in the infected mice. Histopathological investigations revealed visible necrosis of intestinal epithelial cells, shedding of cellular debris in the intestine, while in the kidney, necrosis of renal cortical portion of tubular epithelial cells was observed. STEC O157:H7 is prevalent along PSCP around Hubei of China in different proportions being alarmingly higher in supply chain and markets which is a matter of concern for public health. Copyright © 2017. Published by Elsevier Ltd.

  11. Control of the Biofilms Formed by Curli- and Cellulose-Expressing Shiga Toxin-Producing Escherichia coli Using Treatments with Organic Acids and Commercial Sanitizers.

    Science.gov (United States)

    Park, Yoen Ju; Chen, Jinru

    2015-05-01

    Biofilms are a mixture of bacteria and extracellular products secreted by bacterial cells and are of great concern to the food industry because they offer physical, mechanical, and biological protection to bacterial cells. This study was conducted to quantify biofilms formed by different Shiga toxin-producing Escherichia coli (STEC) strains on polystyrene and stainless steel surfaces and to determine the effectiveness of sanitizing treatments in control of these biofilms. STEC producing various amounts of cellulose (n = 6) or curli (n = 6) were allowed to develop biofilms on polystyrene and stainless steel surfaces at 28°C for 7 days. The biofilms were treated with 2% acetic or lactic acid and manufacturer-recommended concentrations of acidic or alkaline sanitizers, and residual biofilms were quantified. Treatments with the acidic and alkaline sanitizers were more effective than those with the organic acids for removing the biofilms. Compared with their counterparts, cells expressing a greater amount of cellulose or curli formed more biofilm mass and had greater residual mass after sanitizing treatments on polystyrene than on stainless steel. Research suggests that the organic acids and sanitizers used in the present study differed in their ability to control biofilms. Bacterial surface components and cell contact surfaces can influence both biofilm formation and the efficacy of sanitizing treatments. These results provide additional information on control of biofilms formed by STEC.

  12. Sekuen Nukleotida Gene Shiga like toxin-2 dari Isolat Lokal Escherichia coli O157:H7 asal Hewan dan Manusia (NUCLEOTIDES SQUENCES OF SHIGA-LIKE TOXIN 2 GENES OF ESCHERICHIA COLI O157:H7 LOCAL ISOLATES ORIGINATED FROM ANIMALS AND HUMAN

    Directory of Open Access Journals (Sweden)

    I Wayan Suardana

    2017-04-01

    Full Text Available Animals/livestock, especially cattle, are known as the main reservoir of Escherichia coli O157: H7. As the only one of zoonotic E. coli, the pathogenicity of these bacteria is determined by its ability to produce one or more very potent cytotoxin known as Shiga-like toxin (Stx or verocytotoxin, particularly of the Stx2 type that is closely related to the incidence of hemolytic uremic syndrome (HUS in humans. This study analyzed the nucleotide sequences of stx2 gene between isolates from animals and humans in an effort to assess the potential zoonoses of the agent. The research activity was initiated by cultivating 20 isolates of E. coli O157:H7 collection based on result in the previous study i.e. 2 isolates originated from cattle feces, 2 isolates originated from beef, 2 isolates originated from chicken feces, 2 isolates originated from human feces, and 12 non-clinical isolates originated from human fecal who were suffering with renal failure. All isolates were confirmed on selective medium Sorbitol MacConkey Agar (SMAC followed by testing on aglutination O157 latex test, and H7 antisera. Molecular analysis of stx2 gene covering open reading frame (ORF of the stx2 gene was performed using the primer which was designed by researcher i.e. Stx2 (F/Stx2 (R. The results showed, there were 2 isolates i.e. KL-48 (2 originated from human feces and SM-25 (1 originated from cattle feces were positive for carrying a stx2 gene, which was marked by the 1587 bp PCR product. Analysis of sequencing showed both isolates had identical to stx2 nucleotide squences with E. phaga 933 as well as E. coli ATCC 933. These results indicate the both local isolates are potential as zoonotic agents with clinical effects similar to E. phaga 933 and E. coli ATCC 43894. ABSTRAK Hewan ternak khususnya sapi, dikenal sebagai reservoir utama Escherichia coli O157:H7. Sebagai satu-satunya serotipe E. coli yang bersifat zoonosis, patogenitas bakteri ini ditentukan oleh kemampuannya

  13. No evidence of the Shiga toxin-producing E. coli O104:H4 outbreak strain or enteroaggregative E. coli (EAEC found in cattle faeces in northern Germany, the hotspot of the 2011 HUS outbreak area

    Directory of Open Access Journals (Sweden)

    Wieler Lothar H

    2011-11-01

    Full Text Available Abstract Background Ruminants, in particular bovines, are the primary reservoir of Shiga toxin-producing E. coli (STEC, but whole genome analyses of the current German ESBL-producing O104:H4 outbreak strain of sequence type (ST 678 showed this strain to be highly similar to enteroaggregative E. coli (EAEC. Strains of the EAEC pathotype are basically adapted to the human host. To clarify whether in contrast to this paradigm, the O104:H4 outbreak strain and/or EAEC may also be able to colonize ruminants, we screened a total of 2.000 colonies from faecal samples of 100 cattle from 34 different farms - all located in the HUS outbreak region of Northern Germany - for genes associated with the O104:H4 HUS outbreak strain (stx2, terD, rfbO104, fliCH4, STEC (stx1, stx2, escV, EAEC (pAA, aggR, astA, and ESBL-production (blaCTX-M, blaTEM, blaSHV. Results The faecal samples contained neither the HUS outbreak strain nor any EAEC. As the current outbreak strain belongs to ST678 and displays an en-teroaggregative and ESBL-producing phenotype, we additionally screened selected strains for ST678 as well as the aggregative adhesion pattern in HEp-2 cells. However, we were unable to find any strains belonging to ST678 or showing an aggregative adhesion pattern. A high percentage of animals (28% shed STEC, corroborating previous knowl-edge and thereby proving the validity of our study. One of the STEC also harboured the LEE pathogenicity island. In addition, eleven animals shed ESBL-producing E. coli. Conclusions While we are aware of the limitations of our survey, our data support the theory, that, in contrast to other Shiga-toxin producing E. coli, cattle are not the reservoir for the O104:H4 outbreak strain or other EAEC, but that the outbreak strain seems to be adapted to humans or might have yet another reservoir, raising new questions about the epidemiology of STEC O104:H4.

  14. Epidemiology of Shiga toxin producing Escherichia coli in Australia, 2000-2010

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    Vally Hassan

    2012-01-01

    Full Text Available Abstract Background Shiga toxin-producing Escherichia coli (STEC are an important cause of gastroenteritis in Australia and worldwide and can also result in serious sequelae such as haemolytic uraemic syndrome (HUS. In this paper we describe the epidemiology of STEC in Australia using the latest available data. Methods National and state notifications data, as well as data on serotypes, hospitalizations, mortality and outbreaks were examined. Results For the 11 year period 2000 to 2010, the overall annual Australian rate of all notified STEC illness was 0.4 cases per 100,000 per year. In total, there were 822 STEC infections notified in Australia over this period, with a low of 1 notification in the Australian Capital Territory (corresponding to a rate of 0.03 cases per 100,000/year and a high of 413 notifications in South Australia (corresponding to a rate of 2.4 cases per 100,000/year, the state with the most comprehensive surveillance for STEC infection in the country. Nationally, 71.2% (504/708 of STEC infections underwent serotype testing between 2001 and 2009, and of these, 58.0% (225/388 were found to be O157 strains, with O111 (13.7% and O26 (11.1% strains also commonly associated with STEC infections. The notification rate for STEC O157 infections Australia wide between 2001-2009 was 0.12 cases per 100,000 per year. Over the same 9 year period there were 11 outbreaks caused by STEC, with these outbreaks generally being small in size and caused by a variety of serogroups. The overall annual rate of notified HUS in Australia between 2000 and 2010 was 0.07 cases per 100,000 per year. Both STEC infections and HUS cases showed a similar seasonal distribution, with a larger proportion of reported cases occurring in the summer months of December to February. Conclusions STEC infections in Australia have remained fairly steady over the past 11 years. Overall, the incidence and burden of disease due to STEC and HUS in Australia appears

  15. Gene and antigen markers of Shiga-toxin producing E. coli from Michigan and Indiana river water: Occurrence and relation to recreational water quality criteria

    Science.gov (United States)

    Duris, J.W.; Haack, S.K.; Fogarty, L.R.

    2009-01-01

    The relation of bacterial pathogen occurrence to fecal indicator bacteria (FIB) concentrations used for recreational water quality criteria (RWQC) is poorly understood. This study determined the occurrence of Shiga-toxin producing Escherichia coli (STEC) markers and their relation to FIB concentrations in Michigan and Indiana river water. Using 67 fecal coliform (FC) bacteria cultures from 41 river sites in multiple watersheds, we evaluated the occurrence of five STEC markers: the Escherichia coli (EC) O157 antigen and gene, and the STEC virulence genes eaeA, stx1, and stx2. Simple isolations from selected FC cultures yielded viable EC O157. By both antigen and gene assays, EC O157 was detected in a greater proportion of samples exceeding rather than meeting FC RWQC (P public health decision-making. Copyright ?? 2009 by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America. All rights reserved.

  16. Spread and change in stress resistance of Shiga toxin-producing Escherichia coli O157 on fungal colonies.

    Science.gov (United States)

    Lee, Ken-Ichi; Kobayashi, Naoki; Watanabe, Maiko; Sugita-Konishi, Yoshiko; Tsubone, Hirokazu; Kumagai, Susumu; Hara-Kudo, Yukiko

    2014-11-01

    To elucidate the effect of fungal hyphae on the behaviour of Shiga toxin-producing Escherichia coli (STEC) O157, the spread and change in stress resistance of the bacterium were evaluated after coculture with 11 species of food-related fungi including fermentation starters. Spread distances of STEC O157 varied depending on the co-cultured fungal species, and the motile bacterial strain spread for longer distances than the non-motile strain. The population of STEC O157 increased when co-cultured on colonies of nine fungal species but decreased on colonies of Emericella nidulans and Aspergillus ochraceus. Confocal scanning microscopy visualization of green fluorescent protein-tagged STEC O157 on fungal hyphae revealed that the bacterium colonized in the water film that existed on and between hyphae. To investigate the physiological changes in STEC O157 caused by co-culturing with fungi, the bacterium was harvested after 7 days of co-culturing and tested for acid resistance. After co-culture with eight fungal species, STEC O157 showed greater acid resistance compared to those cultured without fungi. Our results indicate that fungal hyphae can spread the contamination of STEC O157 and can also enhance the stress resistance of the bacteria. © 2013 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  17. Assessment of Shiga toxin-producing Escherichia coli isolates from wildlife meat as potential pathogens for humans.

    Science.gov (United States)

    Miko, Angelika; Pries, Karin; Haby, Sabine; Steege, Katja; Albrecht, Nadine; Krause, Gladys; Beutin, Lothar

    2009-10-01

    A total of 140 Shiga toxin-producing Escherichia coli (STEC) strains from wildlife meat (deer, wild boar, and hare) isolated in Germany between 1998 and 2006 were characterized with respect to their serotypes and virulence markers associated with human pathogenicity. The strains grouped into 38 serotypes, but eight O groups (21, 146, 128, 113, 22, 88, 6, and 91) and four H types (21, 28, 2, and 8) accounted for 71.4% and 75.7% of all STEC strains from game, respectively. Eighteen of the serotypes, including enterohemorrhagic E. coli (EHEC) O26:[H11] and O103:H2, were previously found to be associated with human illness. Genes linked to high-level virulence for humans (stx(2), stx(2d), and eae) were present in 46 (32.8%) STEC strains from game. Fifty-four STEC isolates from game belonged to serotypes which are frequently found in human patients (O103:H2, O26:H11, O113:H21, O91:H21, O128:H2, O146:H21, and O146:H28). These 54 STEC isolates were compared with 101 STEC isolates belonging to the same serotypes isolated from farm animals, from their food products, and from human patients. Within a given serotype, most STEC strains were similar with respect to their stx genotypes and other virulence attributes, regardless of origin. The 155 STEC strains were analyzed for genetic similarity by XbaI pulsed-field gel electrophoresis. O103:H2, O26:H11, O113:H21, O128:H2, and O146:H28 STEC isolates from game were 85 to 100% similar to STEC isolates of the same strains from human patients. By multilocus sequence typing, game EHEC O103:H2 strains were attributed to a clonal lineage associated with hemorrhagic diseases in humans. The results from our study indicate that game animals represent a reservoir for and a potential source of human pathogenic STEC and EHEC strains.

  18. Assessment of Shiga Toxin-Producing Escherichia coli Isolates from Wildlife Meat as Potential Pathogens for Humans▿

    Science.gov (United States)

    Miko, Angelika; Pries, Karin; Haby, Sabine; Steege, Katja; Albrecht, Nadine; Krause, Gladys; Beutin, Lothar

    2009-01-01

    A total of 140 Shiga toxin-producing Escherichia coli (STEC) strains from wildlife meat (deer, wild boar, and hare) isolated in Germany between 1998 and 2006 were characterized with respect to their serotypes and virulence markers associated with human pathogenicity. The strains grouped into 38 serotypes, but eight O groups (21, 146, 128, 113, 22, 88, 6, and 91) and four H types (21, 28, 2, and 8) accounted for 71.4% and 75.7% of all STEC strains from game, respectively. Eighteen of the serotypes, including enterohemorrhagic E. coli (EHEC) O26:[H11] and O103:H2, were previously found to be associated with human illness. Genes linked to high-level virulence for humans (stx2, stx2d, and eae) were present in 46 (32.8%) STEC strains from game. Fifty-four STEC isolates from game belonged to serotypes which are frequently found in human patients (O103:H2, O26:H11, O113:H21, O91:H21, O128:H2, O146:H21, and O146:H28). These 54 STEC isolates were compared with 101 STEC isolates belonging to the same serotypes isolated from farm animals, from their food products, and from human patients. Within a given serotype, most STEC strains were similar with respect to their stx genotypes and other virulence attributes, regardless of origin. The 155 STEC strains were analyzed for genetic similarity by XbaI pulsed-field gel electrophoresis. O103:H2, O26:H11, O113:H21, O128:H2, and O146:H28 STEC isolates from game were 85 to 100% similar to STEC isolates of the same strains from human patients. By multilocus sequence typing, game EHEC O103:H2 strains were attributed to a clonal lineage associated with hemorrhagic diseases in humans. The results from our study indicate that game animals represent a reservoir for and a potential source of human pathogenic STEC and EHEC strains. PMID:19700552

  19. House Flies in the Confined Cattle Environment Carry Non-O157 Shiga Toxin-Producing Escherichia coli.

    Science.gov (United States)

    Puri-Giri, R; Ghosh, A; Thomson, J L; Zurek, L

    2017-05-01

    Cattle manure is one of the primary larval developmental habitats of house flies, Musca domestica (L.). Cattle serve as asymptomatic reservoirs of Shiga toxin-producing Escherichia coli (STEC), and bacteria are released into the environment in cattle feces. The USDA-FSIS declared seven STEC serogroups (O157, O26, O45, O103, O145, O121, and O111) as adulterants in beef products. In addition, the serogroup O104 was a culprit of a large outbreak in Germany in 2011. Our study aimed to assess the prevalence of seven non-O157 STEC (O26, O45, O145, O103, O121, O111, and O104) serogroups in adult house flies. Flies (n = 463) were collected from nine feedlots and three dairy farms in six states in the United States and individually processed. This involved a culturing approach with immunomagnetic separation followed by multiplex polymerase chain reactions for detection of individual serogroups and virulence traits. The concentration of bacteria on modified Possé agar ranged between 1.0 × 101 and 7.0 × 107 (mean: 1.5 ± 0.3 × 106) CFU/fly. Out of 463 house flies, 159 (34.3%) carried one or more of six E. coli serogroups of interest. However, STEC was found in 1.5% of house flies from feedlots only. These were E. coli O103 and O104 harboring stx1 and ehxA and E. coli O45 with stx1, eae, and ehxA. This is the first study reporting the isolation of non-O157 STEC in house flies from the confined cattle environment and indicating a potential role of this insect as a vector and reservoir of non-O157 STEC in confined beef cattle. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  20. Enteroaggregative Shiga toxin-producing Escherichia coli of serotype O104:H4 in Belgium and Luxembourg

    Directory of Open Access Journals (Sweden)

    K. De Rauw

    2014-09-01

    Full Text Available In 2011, a large outbreak of infections caused by Shiga toxin-producing Escherichia coli (STEC O104:H4 occurred in Germany. This exceptionally virulent strain combined virulence factors of enteroaggregative E. coli (EAggEC and STEC. After the outbreak only a few sporadic cases of infection with this rare serotype were reported, most of which were related to travel to the Middle East or North Africa. Here we describe two cases of enteroaggregative STEC (Agg-STEC O104:H4 infection that occurred in Belgium in 2012 and 2013 respectively. In both cases travel in a Mediterranean country preceded the infection. The first strain was isolated from the stool of a 42-year-old woman presenting bloody diarrhoea, who had travelled to Tunisia the week before. The second case involves a 14-year-old girl who, upon her return from Turkey to Belgium, suffered from an episode of bloody diarrhoea and haemolytic uraemic syndrome. Extended typing of the isolates with pulsed field gel electrophoresis revealed that the strains were closely related, though not exactly the same as the 2011 outbreak strain. This report supports the previously made hypothesis that Agg-STEC has a human reservoir and might be imported by travellers coming from an area where the pathogen is endemic. Furthermore, it emphasizes the concern that these bacteria may cause future outbreaks as evenly virulent O104:H4 isolates seem to be widespread.

  1. Bacterial community affects toxin production by Gymnodinium catenatum.

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    Maria E Albinsson

    Full Text Available The paralytic shellfish toxin (PST-producing dinoflagellate Gymnodinium catenatum grows in association with a complex marine bacterial community that is both essential for growth and can alter culture growth dynamics. Using a bacterial community replacement approach, we examined the intracellular PST content, production rate, and profile of G. catenatum cultures grown with bacterial communities of differing complexity and composition. Clonal offspring were established from surface-sterilized resting cysts (produced by sexual crosses of strain GCDE06 and strain GCLV01 and grown with: 1 complex bacterial communities derived from each of the two parent cultures; 2 simplified bacterial communities composed of the G. catenatum-associated bacteria Marinobacter sp. strain DG879 or Alcanivorax sp. strain DG881; 3 a complex bacterial community associated with an untreated, unsterilized sexual cross of the parents. Toxin content (STX-equivalent per cell of clonal offspring (134-197 fmol STX cell(-1 was similar to the parent cultures (169-206 fmol STX cell(-1, however cultures grown with single bacterial types contained less toxin (134-146 fmol STX cell(-1 than offspring or parent cultures grown with more complex mixed bacterial communities (152-176 fmol STX cell(-1. Specific toxin production rate (fmol STX day(-1 was strongly correlated with culture growth rate. Net toxin production rate (fmol STX cell(-1 day(-1 did not differ among treatments, however, mean net toxin production rate of offspring was 8-fold lower than the parent cultures, suggesting that completion of the sexual lifecycle in laboratory cultures leads to reduced toxin production. The PST profiles of offspring cultures were most similar to parent GCDE06 with the exception of cultures grown with Marinobacter sp. DG879 which produced higher proportions of dcGTX2+3 and GC1+2, and lower proportions of C1+2 and C3+4. Our data demonstrate that the bacterial community can alter intracellular STX

  2. Bacterial community affects toxin production by Gymnodinium catenatum.

    Science.gov (United States)

    Albinsson, Maria E; Negri, Andrew P; Blackburn, Susan I; Bolch, Christopher J S

    2014-01-01

    The paralytic shellfish toxin (PST)-producing dinoflagellate Gymnodinium catenatum grows in association with a complex marine bacterial community that is both essential for growth and can alter culture growth dynamics. Using a bacterial community replacement approach, we examined the intracellular PST content, production rate, and profile of G. catenatum cultures grown with bacterial communities of differing complexity and composition. Clonal offspring were established from surface-sterilized resting cysts (produced by sexual crosses of strain GCDE06 and strain GCLV01) and grown with: 1) complex bacterial communities derived from each of the two parent cultures; 2) simplified bacterial communities composed of the G. catenatum-associated bacteria Marinobacter sp. strain DG879 or Alcanivorax sp. strain DG881; 3) a complex bacterial community associated with an untreated, unsterilized sexual cross of the parents. Toxin content (STX-equivalent per cell) of clonal offspring (134-197 fmol STX cell(-1)) was similar to the parent cultures (169-206 fmol STX cell(-1)), however cultures grown with single bacterial types contained less toxin (134-146 fmol STX cell(-1)) than offspring or parent cultures grown with more complex mixed bacterial communities (152-176 fmol STX cell(-1)). Specific toxin production rate (fmol STX day(-1)) was strongly correlated with culture growth rate. Net toxin production rate (fmol STX cell(-1) day(-1)) did not differ among treatments, however, mean net toxin production rate of offspring was 8-fold lower than the parent cultures, suggesting that completion of the sexual lifecycle in laboratory cultures leads to reduced toxin production. The PST profiles of offspring cultures were most similar to parent GCDE06 with the exception of cultures grown with Marinobacter sp. DG879 which produced higher proportions of dcGTX2+3 and GC1+2, and lower proportions of C1+2 and C3+4. Our data demonstrate that the bacterial community can alter intracellular STX

  3. Protection of Human Podocytes from Shiga Toxin 2-Induced Phosphorylation of Mitogen-Activated Protein Kinases and Apoptosis by Human Serum Amyloid P Component

    Science.gov (United States)

    Dettmar, Anne K.; Binder, Elisabeth; Greiner, Friederike R.; Liebau, Max C.; Kurschat, Christine E.; Jungraithmayr, Therese C.; Saleem, Moin A.; Schmitt, Claus-Peter; Feifel, Elisabeth; Orth-Höller, Dorothea; Kemper, Markus J.; Pepys, Mark; Würzner, Reinhard

    2014-01-01

    Hemolytic uremic syndrome (HUS) is mainly induced by Shiga toxin 2 (Stx2)-producing Escherichia coli. Proteinuria can occur in the early phase of the disease, and its persistence determines the renal prognosis. Stx2 may injure podocytes and induce proteinuria. Human serum amyloid P component (SAP), a member of the pentraxin family, has been shown to protect against Stx2-induced lethality in mice in vivo, presumably by specific binding to the toxin. We therefore tested the hypothesis that SAP can protect against Stx2-induced injury of human podocytes. To elucidate the mechanisms underlying podocyte injury in HUS-associated proteinuria, we assessed Stx2-induced activation of mitogen-activated protein kinases (MAPKs) and apoptosis in immortalized human podocytes and evaluated the impact of SAP on Stx2-induced damage. Human podocytes express Stx2-binding globotriaosylceramide 3. Stx2 applied to cultured podocytes was internalized and then activated p38α MAPK and c-Jun N-terminal kinase (JNK), important signaling steps in cell differentiation and apoptosis. Stx2 also activated caspase 3, resulting in an increased level of apoptosis. Coincubation of podocytes with SAP and Stx2 mitigated the effects of Stx2 and induced upregulation of antiapoptotic Bcl2. These data suggest that podocytes are a target of Stx2 and that SAP protects podocytes against Stx2-induced injury. SAP may therefore be a useful therapeutic option. PMID:24566618

  4. Molecular screening of bovine raw milk for the presence of Shiga toxin-producing Escherichia coli (STEC on dairy farms

    Directory of Open Access Journals (Sweden)

    Tatiane Vendramin

    2014-09-01

    Full Text Available Milkborne transmission of Shiga toxin- producing Escherichia coli (STEC has raised considerable concern due to recent outbreaks worldwide and poses a threat to public health. The aim of this study was to develop a sensitive and specific multiplex PCR assay to detect the presence of STEC in bovine raw milk. To identify E. coli (ATCC 25922 contamination, the gene uspA was used, and PCR sensitivity and specificity were accessed by testing diluted samples ranging from 2 to 2.0 × 10(6 CFU/mL. To detect STEC, the stx1 and stx2 genes were selected as targets. After reaction standardization, the multiplex assay was tested in raw milk collected from 101 cows on dairy farms. PCR assay for E. coli detection had a specificity of 100% and sensitivity of 79% (P<0.0001, with a lower detection limit of 2 CFU/mL. Multiplex PCR assay had 100% sensitivity for E. coli positive raw milk samples, and 31.1% were contaminated with STEC, 28.3% of stx2, and 1.9% of stx1. The multiplex PCR assay described in the present study can be employed to identify and screen E. coli harboring stx1 and stx2 genes in raw milk on dairy farms and in industries.

  5. Development of a robust method for isolation of shiga toxin-positive Escherichia coli (STEC from fecal, plant, soil and water samples from a leafy greens production region in California.

    Directory of Open Access Journals (Sweden)

    Michael B Cooley

    Full Text Available During a 2.5-year survey of 33 farms and ranches in a major leafy greens production region in California, 13,650 produce, soil, livestock, wildlife, and water samples were tested for Shiga toxin (stx-producing Escherichia coli (STEC. Overall, 357 and 1,912 samples were positive for E. coli O157:H7 (2.6% or non-O157 STEC (14.0%, respectively. Isolates differentiated by O-typing ELISA and multilocus variable number tandem repeat analysis (MLVA resulted in 697 O157:H7 and 3,256 non-O157 STEC isolates saved for further analysis. Cattle (7.1%, feral swine (4.7%, sediment (4.4%, and water (3.3% samples were positive for E. coli O157:H7; 7/32 birds, 2/145 coyotes, 3/88 samples from elk also were positive. Non-O157 STEC were at approximately 5-fold higher incidence compared to O157 STEC: cattle (37.9%, feral swine (21.4%, birds (2.4%, small mammals (3.5%, deer or elk (8.3%, water (14.0%, sediment (12.3%, produce (0.3% and soil adjacent to produce (0.6%. stx1, stx2 and stx1/stx2 genes were detected in 63%, 74% and 35% of STEC isolates, respectively. Subtilase, intimin and hemolysin genes were present in 28%, 25% and 79% of non-O157 STEC, respectively; 23% were of the "Top 6″ O-types. The initial method was modified twice during the study revealing evidence of culture bias based on differences in virulence and O-antigen profiles. MLVA typing revealed a diverse collection of O157 and non-O157 STEC strains isolated from multiple locations and sources and O157 STEC strains matching outbreak strains. These results emphasize the importance of multiple approaches for isolation of non-O157 STEC, that livestock and wildlife are common sources of potentially virulent STEC, and evidence of STEC persistence and movement in a leafy greens production environment.

  6. Toxin production in Dinophysis and the fate of these toxins in marine mussels

    DEFF Research Database (Denmark)

    Nielsen, Lasse Tor

    Diarrhetic shellfish poisoning (DSP) poses a considerable threat to food safety and to the economy of shellfish fishers and farmers in many parts of the world. Thousands of DSP intoxications have been reported, and bivalve harvesting can sometimes be closed down several months in a row. The toxins....... acuta. I grew the two species in laboratory cultures at different irradiances (7-130 μmol photons m-2 s-1) and with different food availability. The results showed that irradiance had no effects on toxin profiles, and only limited effects of the cellular toxin contents. Rather, toxin production rates...... are primarily produced by the marine mixotrophic dinoflagellates Dinophysis spp., known to occur in most parts of the world. Dinophysis can, along with other planktonic organisms, be consumed by filter-feeding bivalves, and thus the toxins can accumulate. Dinophysis can produce the three toxin groups, okadaic...

  7. Soluble CD40 Ligand and Oxidative Response Are Reciprocally Stimulated during Shiga Toxin-Associated Hemolytic Uremic Syndrome

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    Maria J. Abrey Recalde

    2017-10-01

    Full Text Available Shiga toxin (Stx, produced by Escherichia coli, is the main pathogenic factor of diarrhea-associated hemolytic uremic syndrome (HUS, which is characterized by the obstruction of renal microvasculature by platelet-fibrin thrombi. It is well known that the oxidative imbalance generated by Stx induces platelet activation, contributing to thrombus formation. Moreover, activated platelets release soluble CD40 ligand (sCD40L, which in turn contributes to oxidative imbalance, triggering the release of reactive oxidative species (ROS on various cellular types. The aim of this work was to determine if the interaction between the oxidative response and platelet-derived sCD40L, as consequence of Stx-induced endothelium damage, participates in the pathogenic mechanism during HUS. Activated human glomerular endothelial cells (HGEC by Stx2 induced platelets to adhere to them. Although platelet adhesion did not contribute to endothelial damage, high levels of sCD40L were released to the medium. The release of sCD40L by activated platelets was inhibited by antioxidant treatment. Furthermore, we found increased levels of sCD40L in plasma from HUS patients, which were also able to trigger the respiratory burst in monocytes in a sCD40L-dependent manner. Thus, we concluded that platelet-derived sCD40L and the oxidative response are reciprocally stimulated during Stx2-associated HUS. This process may contribute to the evolution of glomerular occlusion and the microangiopathic lesions.

  8. Soluble CD40 Ligand and Oxidative Response Are Reciprocally Stimulated during Shiga Toxin-Associated Hemolytic Uremic Syndrome

    Science.gov (United States)

    Abrey Recalde, Maria J.; Alvarez, Romina S.; Alberto, Fabiana; Mejias, Maria P.; Ramos, Maria V.; Fernandez Brando, Romina J.; Bruballa, Andrea C.; Exeni, Ramon A.; Alconcher, Laura; Ibarra, Cristina A.; Amaral, María M.; Palermo, Marina S.

    2017-01-01

    Shiga toxin (Stx), produced by Escherichia coli, is the main pathogenic factor of diarrhea-associated hemolytic uremic syndrome (HUS), which is characterized by the obstruction of renal microvasculature by platelet-fibrin thrombi. It is well known that the oxidative imbalance generated by Stx induces platelet activation, contributing to thrombus formation. Moreover, activated platelets release soluble CD40 ligand (sCD40L), which in turn contributes to oxidative imbalance, triggering the release of reactive oxidative species (ROS) on various cellular types. The aim of this work was to determine if the interaction between the oxidative response and platelet-derived sCD40L, as consequence of Stx-induced endothelium damage, participates in the pathogenic mechanism during HUS. Activated human glomerular endothelial cells (HGEC) by Stx2 induced platelets to adhere to them. Although platelet adhesion did not contribute to endothelial damage, high levels of sCD40L were released to the medium. The release of sCD40L by activated platelets was inhibited by antioxidant treatment. Furthermore, we found increased levels of sCD40L in plasma from HUS patients, which were also able to trigger the respiratory burst in monocytes in a sCD40L-dependent manner. Thus, we concluded that platelet-derived sCD40L and the oxidative response are reciprocally stimulated during Stx2-associated HUS. This process may contribute to the evolution of glomerular occlusion and the microangiopathic lesions. PMID:29068360

  9. Wild ungulates as disseminators of Shiga toxin-producing Escherichia coli in urban areas.

    Directory of Open Access Journals (Sweden)

    Alan B Franklin

    Full Text Available BACKGROUND: In 2008, children playing on a soccer field in Colorado were sickened with a strain of Shiga toxin-producing Escherichia coli (STEC O157:H7, which was ultimately linked to feces from wild Rocky Mountain elk. We addressed whether wild cervids were a potential source of STEC infections in humans and whether STEC was ubiquitous throughout wild cervid populations in Colorado. METHODOLOGY/PRINCIPAL FINDINGS: We collected 483 fecal samples from Rocky Mountain elk and mule deer in urban and non-urban areas. Samples testing positive for STEC were higher in urban (11.0% than non-urban (1.6% areas. Elk fecal samples in urban areas had a much higher probability of containing STEC, which increased in both urban and non-urban areas as maximum daily temperature increased. Of the STEC-positive samples, 25% contained stx1 strains, 34.3% contained stx2, and 13% contained both stx1 and stx2. Additionally, eaeA genes were detected in 54.1% of the positive samples. Serotypes O103, and O146 were found in elk and deer feces, which also have the potential to cause human illness. CONCLUSIONS/SIGNIFICANCE: The high incidence of stx2 strains combined with eaeA and E-hyl genes that we found in wild cervid feces is associated with severe human disease, such as hemolytic uremic syndrome. This is of concern because there is a very close physical interface between elk and humans in urban areas that we sampled. In addition, we found a strong relationship between ambient temperature and incidence of STEC in elk feces, suggesting a higher incidence of STEC in elk feces in public areas on warmer days, which in turn may increase the likelihood that people will come in contact with infected feces. These concerns also have implications to other urban areas where high densities of coexisting wild cervids and humans interact on a regular basis.

  10. Simple method for Shiga toxin 2e purification by affinity chromatography via binding to the divinyl sulfone group.

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    Hideyuki Arimitsu

    Full Text Available Here we describe a simple affinity purification method for Shiga toxin 2e (Stx2e, a major causative factor of edema disease in swine. Escherichia coli strain MV1184 transformed with the expression plasmid pBSK-Stx2e produced Stx2e when cultivated in CAYE broth containing lincomycin. Stx2e bound to commercial D-galactose gel, containing α-D-galactose immobilized on agarose resin via a divinyl sulfone linker, and was eluted with phosphate-buffered saline containing 4.5 M MgCl2. A small amount of Stx2e bound to another commercial α-galactose-immobilized agarose resin, but not to β-galactose-immobilized resin. In addition, Stx2e bound to thiophilic adsorbent resin containing β-mercaptoethanol immobilized on agarose resin via a divinyl sulfone, and was purified in the same manner as from D-galactose gel, but the Stx2e sample contained some contamination. These results indicate that Stx2e bound to D-galactose gel mainly through the divinyl sulfone group on the resin and to a lesser extent through α-D-galactose. With these methods, the yields of Stx2e and attenuated mutant Stx2e (mStx2e from 1 L of culture were approximately 36 mg and 27.7 mg, respectively, and the binding capacity of the D-galactose gel and thiophilic adsorbent resin for Stx2e was at least 20 mg per 1 ml of resin. In addition, using chimeric toxins with prototype Stx2 which did not bind to thiophilic adsorbent resin and some types of mutant Stx2e and Stx2 which contained inserted mutations in the B subunits, we found that, at the least, asparagine (amino acid 17 of the B subunits was associated with Stx2e binding to the divinyl sulfone group. The mStx2e that was isolated exhibited vaccine effects in ICR mice, indicating that these methods are beneficial for large-scale preparation of Stx2e toxoid, which protects swine from edema disease.

  11. Rapid and simple method by combining FTA™ card DNA extraction with two set multiplex PCR for simultaneous detection of non-O157 Shiga toxin-producing Escherichia coli strains and virulence genes in food samples.

    Science.gov (United States)

    Kim, S A; Park, S H; Lee, S I; Ricke, S C

    2017-12-01

    The aim of this research was to optimize two multiplex polymerase chain reaction (PCR) assays that could simultaneously detect six non-O157 Shiga toxin-producing Escherichia coli (STEC) as well as the three virulence genes. We also investigated the potential of combining the FTA™ card-based DNA extraction with the multiplex PCR assays. Two multiplex PCR assays were optimized using six primer pairs for each non-O157 STEC serogroup and three primer pairs for virulence genes respectively. Each STEC strain specific primer pair only amplified 155, 238, 321, 438, 587 and 750 bp product for O26, O45, O103, O111, O121 and O145 respectively. Three virulence genes were successfully multiplexed: 375 bp for eae, 655 bp for stx1 and 477 bp for stx2. When two multiplex PCR assays were validated with ground beef samples, distinctive bands were also successfully produced. Since the two multiplex PCR examined here can be conducted under the same PCR conditions, the six non-O157 STEC and their virulence genes could be concurrently detected with one run on the thermocycler. In addition, all bands clearly appeared to be amplified by FTA card DNA extraction in the multiplex PCR assay from the ground beef sample, suggesting that an FTA card could be a viable sampling approach for rapid and simple DNA extraction to reduce time and labour and therefore may have practical use for the food industry. Two multiplex polymerase chain reaction (PCR) assays were optimized for discrimination of six non-O157 Shiga toxin-producing Escherichia coli (STEC) and identification of their major virulence genes within a single reaction, simultaneously. This study also determined the successful ability of the FTA™ card as an alternative to commercial DNA extraction method for conducting multiplex STEC PCR assays. The FTA™ card combined with multiplex PCR holds promise for the food industry by offering a simple and rapid DNA sample method for reducing time, cost and labour for detection of STEC in

  12. Role of climate in the spread of shiga toxin-producing Escherichia coli infection among children

    Science.gov (United States)

    Acquaotta, Fiorella; Ardissino, Gianluigi; Fratianni, Simona; Perrone, Michela

    2017-09-01

    Haemolytic-uraemic syndrome (HUS) is a rare disease mainly affecting children that develops as a complication of shiga toxin-producing Escherichia coli (STEC) infection. It is characterised by acute kidney injury, platelet consumption and mechanical destruction of red blood cells (haemolysis). In order to test the working hypothesis that the spread of the infection is influenced by specific climatic conditions, we analysed all of the identified cases of infection occurring between June 2010 and December 2013 in four provinces of Lombardy, Italy (Milano, Monza Brianza, Varese and Brescia), in which a STEC surveillance system has been developed as part of a preventive programme. In the selected provinces, we recorded in few days a great number of cases and clusters which are unrelated for spatially distant or for the disease are caused by different STEC serotypes. In order to investigate a common factor that favoured the onset of infection, we have analysed in detail the weather conditions of the areas. The daily series of temperature, rain and relative humidity were studied to show the common climate peculiarities whilst the correlation coefficient and the principal component analysis (PCA) were used to point out the meteorological variable, maximum temperature, as the principal climate element in the onset of the infection. The use of distributed lag non-linear models (DLNM) and the climate indices characterising heat waves (HWs) has allowed to identify the weather conditions associated with STEC infection. The study highlighted a close temporal correlation between STEC infection in children and the number, duration and frequency of heat waves. In particular, if the maximum temperature is greater than 90th percentile, days classified as very hot, for 3 or more consecutive days, the risk of infection is increasing.

  13. Prevalence of Shiga toxin Producing Escherichia coli, Salmonella enterica and Listeria monocytogenes at Public Access Watershed Sites in a California Central Coast Agricultural Region

    Directory of Open Access Journals (Sweden)

    Michael B Cooley

    2014-03-01

    Full Text Available Produce contaminated with enteric pathogens is a major source of foodborne illness in the United States. Lakes, streams, rivers, and ponds were sampled with Moore swabs bi-monthly for over two years at 30 locations in the vicinity of a leafy green growing region on the Central California Coast and screened for Shiga toxin producing Escherichia coli (STEC, Salmonella enterica, and Listeria monocytogenes to evaluate the prevalence and persistence of pathogen subtypes. The prevalence of STEC from 1,386 samples was 11%; 110 samples (8% contained E. coli O157:H7 with the highest prevalence occurring close to cattle operations. Non-O157 STEC isolates represented major clinical O-types and 57% contained both shiga toxin types 1 and 2 and intimin. Multiple Locus Variable Number Tandem Repeat Analysis of STEC isolates indicated prevalent strains during the period of study. Notably, Salmonella was present at high levels throughout the sampling region with 65% prevalence in 1,405 samples resulting in 996 isolates with slightly lower prevalence in late autumn. There were 2, 8 and 14 sites that were Salmonella-positive over 90%, 80% and 70% of the time, respectively. The serotypes identified most often were 6,8:d:-, Typhimurium, and Give. Interestingly, analysis by Pulsed Field Gel Electrophoresis indicated persistence and transport of pulsotypes in the region over several years. In this original study of L. monocytogenes in the region prevalence was 43% of 1,405 samples resulting in 635 individual isolates. Over 85% of the isolates belonged to serotype 4b with serotypes 1/2a, 1/2b, 3a, 4d with 4e representing the rest, and there were 12 and 2 sites that were positive over 50% and 80% of the time, respectively. Although surface water is not directly used for irrigation in this region, transport to the produce can occur by other means. This environmental survey assesses initial contamination levels towards an understanding of transport leading to produce

  14. Shiga Toxin-Producing Escherichia coli in Plateau Pika (Ochotona curzoniae) on the Qinghai-Tibetan Plateau, China.

    Science.gov (United States)

    Bai, Xiangning; Zhang, Wang; Tang, Xinyuan; Xin, Youquan; Xu, Yanmei; Sun, Hui; Luo, Xuelian; Pu, Ji; Xu, Jianguo; Xiong, Yanwen; Lu, Shan

    2016-01-01

    Shiga toxin-producing Escherichia coli (STEC) are an emerging group of zoonotic pathogens. Ruminants are the natural reservoir of STEC. In this study we determined the prevalence and characteristics of the STEC in plateau pika (Ochotona curzoniae) on the Qinghai-Tibetan Plateau, China. A total of 1116 pika samples, including 294 intestinal contents samples, 317 fecal samples, and 505 intestinal contents samples, were collected from May to August in the years 2012, 2013, and 2015, respectively. Twenty-one samples (1.88%) yielded at least one STEC isolate; in total, 22 STEC isolates were recovered. Thirteen different O serogroups and 14 serotypes were identified. One stx 1 subtype (stx 1a) and three stx 2 subtypes (stx 2a, stx 2b, and stx 2d) were present in the STEC isolates. Fifteen, fourteen, and three STEC isolates harbored the virulence genes ehxA, subA, and astA, respectively. Adherence-associated genes iha and saa were, respectively, present in 72.73 and 68.18% of the STEC isolates. Twenty antibiotics were active against all the STEC isolates; all strains were resistant to penicillin G, and some to cephalothin or streptomycin. The 22 STEC isolates were divided into 16 pulsed-field gel electrophoresis patterns and 12 sequence types. Plateau pikas may play a role in the ongoing circulation of STEC in the Qinghai-Tibetan plateau. This study provides the first report on STEC in plateau pikas and new information about STEC reservoirs in wildlife. Based on the serotypes, virulence gene profiles and multi-locus sequence typing (MLST) analysis, the majority of these pika STECs may pose a low public health risk.

  15. A Topographical Atlas of Shiga Toxin 2e Receptor Distribution in the Tissues of Weaned Piglets

    Directory of Open Access Journals (Sweden)

    Daniel Steil

    2016-11-01

    Full Text Available Shiga toxin (Stx 2e of Stx-producing Escherichia coli (STEC is the primary virulence factor in the development of pig edema disease shortly after weaning. Stx2e binds to the globo-series glycosphingolipids (GSLs globotriaosylceramide (Gb3Cer, Galα1-4Galβ1-4Glcβ1-1Cer and globotetraosylceramide (Gb4Cer, GalNAcβ1-3Galα1-4Galβ1-4Glcβ1-1Cer, the latter acting as the preferential Stx2e receptor. We determined Stx receptor profiles of 25 different tissues of a male and a female weaned piglet using immunochemical solid phase binding assays combined with mass spectrometry. All probed tissues harbored GSL receptors, ranging from high (category I over moderate (category II to low content (category III. Examples of Gb4Cer expression in category I tissues are small intestinal ileum, kidney pelvis and whole blood, followed by colon, small intestinal duodenum and jejunum belonging to category II, and kidney cortex, cerebrum and cerebellum as members of category III organs holding true for both genders. Dominant Gb3Cer and Gb4Cer lipoforms were those with ceramides carrying constant sphingosine (d18:1 and a variable C16:0, C22:0 or C24:1/C24:0 fatty acid. From the mapping data, we created a topographical atlas for Stx2e receptors in piglet tissues and organs, which might be helpful to further investigations on the molecular and cellular mechanisms that underlie infections of Stx2e-producing STEC in pigs and their zoonotic potential for humans.

  16. Shiga toxin-converting phages and the emergence of new pathogenic Escherichia coli: a world in motion

    Science.gov (United States)

    Tozzoli, Rosangela; Grande, Laura; Michelacci, Valeria; Ranieri, Paola; Maugliani, Antonella; Caprioli, Alfredo; Morabito, Stefano

    2014-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) are pathogenic E. coli causing diarrhea, hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). STEC are characterized by a constellation of virulence factors additional to Stx and have long been regarded as capable to cause HC and HUS when possessing the ability of inducing the attaching and effacing (A/E) lesion to the enterocyte, although strains isolated from such severe infections sometimes lack this virulence feature. Interestingly, the capability to cause the A/E lesion is shared with another E. coli pathogroup, the Enteropathogenic E. coli (EPEC). In the very recent times, a different type of STEC broke the scene causing a shift in the paradigm for HUS-associated STEC. In 2011, a STEC O104:H4 caused a large outbreak with more than 800 HUS and 50 deaths. Such a strain presented the adhesion determinants of Enteroaggregative E. coli (EAggEC). We investigated the possibility that, besides STEC and EAggEC, other pathogenic E. coli could be susceptible to infection with stx-phages. A panel of stx2-phages obtained from STEC isolated from human disease was used to infect experimentally E. coli strains representing all the known pathogenic types, including both diarrheagenic E. coli (DEC) and extra-intestinal pathogenic E. coli (ExPEC). We observed that all the E. coli pathogroups used in the infection experiments were susceptible to the infection. Our results suggest that the stx2-phages used may not have specificity for E. coli adapted to the intestinal environment, at least in the conditions used. Additionally, we could only observe transient lysogens suggesting that the event of stable stx2-phage acquisition occurs rarely. PMID:24999453

  17. Short-term evolution of Shiga toxin-producing Escherichia coli O157:H7 between two food-borne outbreaks.

    Science.gov (United States)

    Cowley, Lauren A; Dallman, Timothy J; Fitzgerald, Stephen; Irvine, Neil; Rooney, Paul J; McAteer, Sean P; Day, Martin; Perry, Neil T; Bono, James L; Jenkins, Claire; Gally, David L

    2016-09-01

    Shiga toxin-producing Escherichia coli (STEC) O157:H7 is a public health threat and outbreaks occur worldwide. Here, we investigate genomic differences between related STEC O157:H7 that caused two outbreaks, eight weeks apart, at the same restaurant. Short-read genome sequencing divided the outbreak strains into two sub-clusters separated by only three single-nucleotide polymorphisms in the core genome while traditional typing identified them as separate phage types, PT8 and PT54. Isolates did not cluster with local strains but with those associated with foreign travel to the Middle East/North Africa. Combined long-read sequencing approaches and optical mapping revealed that the two outbreak strains had undergone significant microevolution in the accessory genome with prophage gain, loss and recombination. In addition, the PT54 sub-type had acquired a 240 kbp multi-drug resistance (MDR) IncHI2 plasmid responsible for the phage type switch. A PT54 isolate had a general fitness advantage over a PT8 isolate in rich medium, including an increased capacity to use specific amino acids and dipeptides as a nitrogen source. The second outbreak was considerably larger and there were multiple secondary cases indicative of effective human-to-human transmission. We speculate that MDR plasmid acquisition and prophage changes have adapted the PT54 strain for human infection and transmission. Our study shows the added insights provided by combining whole-genome sequencing approaches for outbreak investigations.

  18. Characterization of enteropathogenic and Shiga toxin-producing Escherichia coli in cattle and deer in a shared agroecosystem

    Directory of Open Access Journals (Sweden)

    Pallavi eSingh

    2015-04-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC is an important foodborne pathogen. Cattle are suggested to be an important reservoir for STEC; however, these pathogens have also been isolated from other livestock and wildlife. In this study we sought to investigate transmission of STEC, enterohemorrhagic E. coli (EHEC and enteropathogenic E. coli (EPEC between cattle and white-tailed deer in a shared agroecosystem. Cattle feces were collected from 100 animals in a Michigan dairy farm in July 2012, while 163 deer fecal samples were collected during two sampling periods (March and June. The locations of deer fecal pellets were recorded via geographic information system mapping and microsatellite multi-locus genotyping was used to link the fecal samples to individual deer at both time points. Following subculture to sorbitol MacConkey agar and STEC CHROMagar, the pathogens were characterized by serotyping, stx profiling, and PCR-based fingerprinting; multilocus sequence typing (MLST was performed on a subset. STEC and EHEC were cultured from 12% and 16% of cattle, respectively, and EPEC was found in 36%. Deer were significantly less likely to have a pathogen in March versus June where the frequency of STEC, EHEC, and EPEC was 1%, 6% and 22%, respectively. PCR fingerprinting and MLST clustered the cattle- and deer-derived strains together in a phylogenetic tree. Two STEC strains recovered from both animal species shared MLST and fingerprinting profiles, thereby providing evidence of interspecies transmission and highlighting the importance of wildlife species in pathogen shedding dynamics and persistence in the environment and cattle herds.

  19. Growth and Survival of Acid-Resistant and Non-Acid-Resistant Shiga-Toxin-Producing Escherichia coli Strains during the Manufacture and Ripening of Camembert Cheese.

    Science.gov (United States)

    Montet, M P; Jamet, E; Ganet, S; Dizin, M; Miszczycha, S; Dunière, L; Thevenot, D; Vernozy-Rozand, C

    2009-01-01

    Growth and survival of acid-resistant (AR) and non-acid-resistant (NAR) Shiga-toxin-producing Escherichia coli (STEC) strains were investigated during the manufacture and ripening of microfiltered milk Camembert cheeses. The induction of acid resistance of the STEC strains in cheeses was also studied. Six different mixtures of AR and/or NAR STEC strains were inoculated separately into microfiltered milk at a level of 10(3) CFU mL(-1). The STEC counts (AR and NAR) initially increased by 1 to 2 log(10) CFU g(-1) during cheese-making. Thereafter, the populations stabilized during salting/drying and then decreased during the early stages of ripening. Exposing the STEC strains in artificially inoculated cheeses to simulated gastric fluid (SGF - pH: 2.0) reduced the number of NAR strains to undetectable levels within 40 minutes, versus 120 minutes for the AR STEC strains. AR and NAR STEC were able to survive during the manufacture and ripening of Camembert cheese prepared from microfiltered milk with no evidence of induced acid tolerance in NAR STEC strains.

  20. Growth and Survival of Acid-Resistant and Non-Acid-Resistant Shiga-Toxin-Producing Escherichia coli Strains during the Manufacture and Ripening of Camembert Cheese

    Directory of Open Access Journals (Sweden)

    M. P. Montet

    2009-01-01

    Full Text Available Growth and survival of acid-resistant (AR and non-acid-resistant (NAR Shiga-toxin-producing Escherichia coli (STEC strains were investigated during the manufacture and ripening of microfiltered milk Camembert cheeses. The induction of acid resistance of the STEC strains in cheeses was also studied. Six different mixtures of AR and/or NAR STEC strains were inoculated separately into microfiltered milk at a level of 103 CFU mL−1. The STEC counts (AR and NAR initially increased by 1 to 2 log⁡10 CFU g−1 during cheese-making. Thereafter, the populations stabilized during salting/drying and then decreased during the early stages of ripening. Exposing the STEC strains in artificially inoculated cheeses to simulated gastric fluid (SGF - pH: 2.0 reduced the number of NAR strains to undetectable levels within 40 minutes, versus 120 minutes for the AR STEC strains. AR and NAR STEC were able to survive during the manufacture and ripening of Camembert cheese prepared from microfiltered milk with no evidence of induced acid tolerance in NAR STEC strains.

  1. In vivo characterization of fusion protein comprising of A1 subunit of Shiga toxin and human GM-CSF: Assessment of its immunogenicity and toxicity.

    Science.gov (United States)

    Oloomi, Mana; Bouzari, Saeid; Shariati, Elaheh

    2010-10-01

    Most cancer cells become resistant to anti-cancer agents. In the last few years, a new approach for targeted therapy of human cancer has been developed using immunotoxins which comprise both the cell targeting and the cell killing moieties. In the present study, the recombinant Shiga toxin A1 subunit fused to human granulocyte-macrophage colony stimulating factor (A1-GM-CSF), previously produced in E. coli, was further characterized. The recombinant protein could cause 50% cytotoxicity and induced apoptosis in cells bearing GM-CSF receptors. The non-specific toxicity of the fusion protein was assessed in C57BL/6 and BALB/c mice. No mortality was observed in either group of mice, with different concentration of fusion protein. The lymphocyte proliferation assay, induction of specific IgG response and a mixed (Th1/Th2) response were observed only in BALB/c mice. The mixed response in BALB/c mice (Th1/Th2) could be explained on the basis of the two components of the fusion protein i.e. A1 and GM-CSF.

  2. Multiplex real-time PCR assays for detection of eight Shiga toxin-producing Escherichia coli in food samples by melting curve analysis.

    Science.gov (United States)

    Singh, Prashant; Mustapha, Azlin

    2015-12-23

    Shiga toxin-producing Escherichia coli (STEC) are pathogenic strains of E. coli that can cause bloody diarrhea and kidney failure. Seven STEC serogroups, O157, O26, O45, O103, O111, O121 and O145 are responsible for more than 71% of the total infections caused by this group of pathogens. All seven serogroups are currently considered as adulterants in non-intact beef products in the U.S. In this study, two multiplex melt curve real-time PCR assays with internal amplification controls (IACs) were standardized for the detection of eight STEC serogroups. The first multiplex assay targeted E. coli serogroups O145, O121, O104, and O157; while the second set detected E. coli serogroups O26, O45, O103 and O111. The applicability of the assays was tested using 11 different meat and produce samples. For food samples spiked with a cocktail of four STEC serogroups with a combined count of 10 CFU/25 g food, all targets of the multiplex assays were detected after an enrichment period of 6h. The assays also worked efficiently when 325 g of food samples were spiked with 10 CFU of STECs. The assays are not dependent on fluorescent-labeled probes or immunomagnetic beads, and can be used for the detection of eight STEC serogroups in less than 11h. Routine preliminary screening of STECs in food samples is performed by testing for the presence of STEC virulence genes. The assays developed in this study can be useful as a first- or second-tier test for the identification of the eight O serogroup-specific genes in suspected food samples. Copyright © 2015. Published by Elsevier B.V.

  3. Influence of Selenium on the Production of T-2 Toxin by Fusarium poae.

    Science.gov (United States)

    Cheng, Bolun; Zhang, Yan; Tong, Bei; Yin, Hong

    2017-07-01

    The objective of this study was to investigate the effects of selenium on the production of T-2 toxin by a Fusarium poae strain cultured in a synthetic medium containing different concentrations of selenium. The T-2 toxin contents in fermentative products were evaluated by a high performance liquid chromatography (HPLC). The results showed that the production of T-2 toxin was correlated with the concentration of selenium added to the medium. In all three treatments, the addition of 1 mg/L selenium to the medium resulted in a lower toxin yield than the control (0 mg/L); the yield of the toxin began to increase when selenium concentration was 10 mg/L, while it decreased again at 20 mg/L. In summary, T-2 toxin yield in the fermentative product was affected by the addition of selenium to the medium, and a selenium concentration of 20 mg/L produced the maximum inhibitory effect of T-2 toxin yield in the fermentative product of F. poae.

  4. Natural killer T (NKT cells accelerate Shiga toxin type 2 (Stx2 pathology in mice

    Directory of Open Access Journals (Sweden)

    Fumiko eObata

    2015-04-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC is a leading cause of childhood renal disease He-molytic Uremic Syndrome (HUS. The involvement of renal cytokines and chemokines is sus-pected to play a critical role in disease progression. In current article, we tested the hypothesis that NKT cells are involved in Stx2-induced pathology in vivo. To address this hypothesis we compared Stx2 toxicity in WT and CD1 knockout (KO mice. In CD1KO mice, which lack nat-ural killer T (NKT cells, Stx2-induced pathologies such as weight loss, renal failure, and death were delayed. In WT mice, Stx2-specific selective increase in urinary albumin occurs in later time points, and this was also delayed in NKT cell deficient mice. NKT cell-associated cytokines such as IL-2, IL-4, IFN-γ and IL-17 were detected in kidney lysates of Stx2-injected WT mice with the peak around 36 h after Stx2 injection. In CD1KO, there was a delay in the kinetics, and increases in these cytokines were observed 60 h post Stx2 injection. These data suggest that NKT cells accelerate Stx2-induced pathology in mouse kidneys. To determine the mechanism by which NKT cells promote Stx2-associated disease, in vitro studies were performed using murine renal cells. We found that murine glomerular endothelial cells and podocytes express functional CD1d molecules and can present exogenous antigen to NKT cells. Moreover, we observed the direct interaction between Stx2 and the receptor Gb3 on the surface of mouse renal cells by 3D STORM-TIRF which provides single molecule imaging. Collectively, these data suggest that Stx2 binds to Gb3 on renal cells and leads to aberrant CD1d-mediated NKT cell activation. Therefore, strategies targeting NKT cells could have a significant impact on Stx2-associated renal pathology in STEC disease.

  5. Natural killer T (NKT) cells accelerate Shiga toxin type 2 (Stx2) pathology in mice.

    Science.gov (United States)

    Obata, Fumiko; Subrahmanyam, Priyanka B; Vozenilek, Aimee E; Hippler, Lauren M; Jeffers, Tynae; Tongsuk, Methinee; Tiper, Irina; Saha, Progyaparamita; Jandhyala, Dakshina M; Kolling, Glynis L; Latinovic, Olga; Webb, Tonya J

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) is a leading cause of childhood renal disease Hemolytic Uremic Syndrome (HUS). The involvement of renal cytokines and chemokines is suspected to play a critical role in disease progression. In current article, we tested the hypothesis that NKT cells are involved in Stx2-induced pathology in vivo. To address this hypothesis we compared Stx2 toxicity in WT and CD1 knockout (KO) mice. In CD1KO mice, which lack natural killer T (NKT) cells, Stx2-induced pathologies such as weight loss, renal failure, and death were delayed. In WT mice, Stx2-specific selective increase in urinary albumin occurs in later time points, and this was also delayed in NKT cell deficient mice. NKT cell-associated cytokines such as IL-2, IL-4, IFN-γ, and IL-17 were detected in kidney lysates of Stx2-injected WT mice with the peak around 36 h after Stx2 injection. In CD1KO, there was a delay in the kinetics, and increases in these cytokines were observed 60 h post Stx2 injection. These data suggest that NKT cells accelerate Stx2-induced pathology in mouse kidneys. To determine the mechanism by which NKT cells promote Stx2-associated disease, in vitro studies were performed using murine renal cells. We found that murine glomerular endothelial cells and podocytes express functional CD1d molecules and can present exogenous antigen to NKT cells. Moreover, we observed the direct interaction between Stx2 and the receptor Gb3 on the surface of mouse renal cells by 3D STORM-TIRF which provides single molecule imaging. Collectively, these data suggest that Stx2 binds to Gb3 on renal cells and leads to aberrant CD1d-mediated NKT cell activation. Therefore, strategies targeting NKT cells could have a significant impact on Stx2-associated renal pathology in STEC disease.

  6. A Review of Current Research and Knowledge Gaps in the Epidemiology of Shiga Toxin-Producing Escherichia coli and Salmonella spp. in Trinidad and Tobago.

    Science.gov (United States)

    Persad, Anil K; LeJeune, Jeffrey

    2018-04-17

    Salmonella and Shiga toxin-producing Escherichia coli are two of the main causes of foodborne disease globally, and while they have been implicated as possible causes of foodborne disease within the Caribbean region, the actual incidence is unknown. Trinidad and Tobago, one of the larger countries in the Caribbean, has an estimated annual foodborne disease burden of over 100,000 cases and, similar to other countries, the etiology of most of these cases is unknown. Both pathogens can reside as part of the normal gastrointestinal microflora of many wild and domestic animals, with animals acting as reservoirs, spillover hosts, or dead-end hosts. Carriage in animal species can be asymptomatic or, in the case of Salmonella in particular, there may be clinical manifestation in animals, which resemble the disease seen in humans. In this review, we will focus on the epidemiology of these two foodborne pathogens in Trinidad and Tobago and identify any knowledge gaps in the published literature. The filling of this critical knowledge void is essential for the development and implementation of appropriate mechanisms to reduce the dissemination and transmission of these pathogens, not only in Trinidad and Tobago, but also in the wider Caribbean.

  7. Real-time genomic investigation underlying the public health response to a Shiga toxin-producing Escherichia coli O26:H11 outbreak in a nursery.

    Science.gov (United States)

    Moran-Gilad, J; Rokney, A; Danino, D; Ferdous, M; Alsana, F; Baum, M; Dukhan, L; Agmon, V; Anuka, E; Valinsky, L; Yishay, R; Grotto, I; Rossen, J W A; Gdalevich, M

    2017-10-01

    Shiga toxin-producing Escherichia coli (STEC) is a significant cause of gastrointestinal infection and the haemolytic-uremic syndrome (HUS). STEC outbreaks are commonly associated with food but animal contact is increasingly being implicated in its transmission. We report an outbreak of STEC affecting young infants at a nursery in a rural community (three HUS cases, one definite case, one probable case, three possible cases and five carriers, based on the combination of clinical, epidemiological and laboratory data) identified using culture-based and molecular techniques. The investigation identified repeated animal contact (animal farming and petting) as a likely source of STEC introduction followed by horizontal transmission. Whole genome sequencing (WGS) was used for real-time investigation of the incident and revealed a unique strain of STEC O26:H11 carrying stx2a and intimin. Following a public health intervention, no additional cases have occurred. This is the first STEC outbreak reported from Israel. WGS proved as a useful tool for rapid laboratory characterization and typing of the outbreak strain and informed the public health response at an early stage of this unusual outbreak.

  8. Expression and purification of recombinant Shiga toxin 2B from ...

    African Journals Online (AJOL)

    sunny t

    2016-05-25

    May 25, 2016 ... MATERIALS AND METHODS. Bacterial strains, plasmid and media ... toxin 2B gene after purified by wizard genomic DNA purification kit. (Promega, USA) ..... This result was approximately two times higher compared to Halo .... manual, 3rd Eds. New York: Cold Spring Harbor Laboratory Press,. Cold Spring ...

  9. Circulating microRNAs in patients with Shiga-Toxin-producing E. coli O104:H4 induced hemolytic uremic syndrome.

    Directory of Open Access Journals (Sweden)

    Johan M Lorenzen

    Full Text Available BACKGROUND: In early May 2011, an outbreak of hemorrhagic colitis associated with hemolytic-uremic syndrome (HUS first developed in Northern Germany and spread to 15 other countries in Europe. The outbreak-strain O104:H4, which combined virulence factors of typical enteroaggregative and Shiga-Toxin-producing E. coli was associated with an unusual high rate of hemolytic uremic syndrome. Also an unexpected high rate of coma and seizures leading to mechanical ventilation and ICU treatment was observed. MicroRNAs are small ribonucleotides orchestrating gene expression. We tested whether circulating microRNAs in serum of HUS patients during the 2011 epidemics are altered in this patient cohort and related to clinical manifestations. METHODOLOGY/PRINCIPAL FINDINGS: We profiled microRNAs using RNA isolated from serum of patients and healthy age-matched controls. The results were validated in 38 patients at baseline, 29 patients during follow-up and 21 age-matched healthy controls by miRNA-specific quantitative RT-PCR. Circulating levels of miR-24, miR-126 were increased in HUS patients versus controls. There was no association between these microRNAs and renal function or the need for renal replacement therapy. In contrast, levels of miR-126 were associated with neurological symptoms at baseline and during follow-up. In addition, miR-126 (on admission and miR-24 (on admission and during follow-up were associated with platelet count. CONCLUSIONS/SIGNIFICANCE: Circulating microRNAs are strongly altered in this patient cohort and associated with neurological symptoms as well as platelet count.

  10. Real-time PCR and enzyme-linked fluorescent assay methods for detecting Shiga-toxin-producing Escherichia coli in mincemeat samples.

    Science.gov (United States)

    Stefan, A; Scaramagli, S; Bergami, R; Mazzini, C; Barbanera, M; Perelle, S; Fach, P

    2007-03-01

    This work aimed to compare real-time polymerase chain reaction (PCR) with the commercially available enzyme-linked fluorescent assay (ELFA) VIDAS ECOLI O157 for detecting Escherichia coli O157 in mincemeat. In addition, a PCR-based survey on Shiga-toxin-producing E. coli (STEC) in mincemeat collected in Italy is presented. Real-time PCR assays targeting the stx genes and a specific STEC O157 sequence (SILO157, a small inserted locus of STEC O157) were tested for their sensitivity on spiked mincemeat samples. After overnight enrichment, the presence of STEC cells could be clearly determined in the 25 g samples containing 10 bacterial cells, while the addition of five bacteria provided equivocal PCR results with Ct values very close to or above the threshold of 40. The PCR tests proved to be more sensitive than the ELFA-VIDAS ECOLI O157, whose detection level started from 50 bacterial cells/25 g of mincemeat. The occurrence of STEC in 106 mincemeat (bovine, veal) samples collected from September to November 2004 at five different points of sale in Italy (one point of sale in Arezzo, Tuscany, central Italy, two in Mantova, Lombardy, Northern Italy, and two in Bologna, Emilia-Romagna, upper-central Italy) was less than 1%. Contamination by the main STEC O-serogroups representing a major public health concern, including O26, O91, O111, O145, and O157, was not detected. This survey indicates that STEC present in these samples are probably not associated with pathogenesis in humans.

  11. Surveillance of Virulence Markers and Antibiotic Resistance of Shiga toxin Producing E.coli O157:H7 Strains from Meats Purchase in Shiraz

    Directory of Open Access Journals (Sweden)

    Mohammad Kargar

    2011-09-01

    Full Text Available Background: Shiga toxin Producing Escherichia coli O157:H7 is a common pathogen in cattle, which occasional causes some human disease. This bacterium can potentially contaminate meat and clinical cases of E.coli O157:H7 infections are often associated with consumption of undercooked ground beef. Methods: In this cross-sectional study 122 samples of ground meat were collected and after enrichment in specific culture media and evaluation sorbitol fermentation and their β-glucoronidase activity, the isolation of E.coli O157:H7 strains have been confirmed with specific antisera. Then virulence genes verotoxin, intimin and hemolysin with multiplex PCR and antibiotic resistance strains with disk diffusion method have been tested. Results: Out of specimens that have been supplied, 119 sorbitol negative colonies isolated which 3 strains O157:H7 (2.45% with specific antisera confirmed. Out of considered virulence genes, in two cases of these samples (1.64% the stx1 and eaeA genes were seen and also 2 isolated bacteria had resistance to erythromycin, tetracycline, ampicillin, penicillin, clindamicin, cefixime, novobiocin, and gentamicin antibiotics. Conclusion: As this organism lives in intestines of healthy cattle, preventive measures on cattle farms and during meat processing are necessary.

  12. Effect of gamma radiation on the reduction of Salmonella strains, Listeria monocytogenes, and Shiga toxin-producing Escherichia coli and sensory evaluation of minimally processed spinach (Tetragonia expansa).

    Science.gov (United States)

    Rezende, Ana Carolina B; Igarashi, Maria Crystina; Destro, Maria Teresa; Franco, Bernadette D G M; Landgraf, Mariza

    2014-10-01

    This study evaluated the effects of irradiation on the reduction of Shiga toxin-producing Escherichia coli (STEC), Salmonella strains, and Listeria monocytogenes, as well as on the sensory characteristics of minimally processed spinach. Spinach samples were inoculated with a cocktail of three strains each of STEC, Salmonella strains, and L. monocytogenes, separately, and were exposed to gamma radiation doses of 0, 0.2, 0.4, 0.6, 0.8, and 1.0 kGy. Samples that were exposed to 0.0, 1.0, and 1.5 kGy and kept under refrigeration (4°C) for 12 days were submitted to sensory analysis. D10 -values ranged from 0.19 to 0.20 kGy for Salmonella and from 0.20 to 0.21 for L. monocytogenes; for STEC, the value was 0.17 kGy. Spinach showed good acceptability, even after exposure to 1.5 kGy. Because gamma radiation reduced the selected pathogens without causing significant changes in the quality of spinach leaves, it may be a useful method to improve safety in the fresh produce industry.

  13. Association of clustered regularly interspaced short palindromic repeat (CRISPR) elements with specific serotypes and virulence potential of shiga toxin-producing Escherichia coli.

    Science.gov (United States)

    Toro, Magaly; Cao, Guojie; Ju, Wenting; Allard, Marc; Barrangou, Rodolphe; Zhao, Shaohua; Brown, Eric; Meng, Jianghong

    2014-02-01

    Shiga toxin-producing Escherichia coli (STEC) strains (n = 194) representing 43 serotypes and E. coli K-12 were examined for clustered regularly interspaced short palindromic repeat (CRISPR) arrays to study genetic relatedness among STEC serotypes. A subset of the strains (n = 81) was further analyzed for subtype I-E cas and virulence genes to determine a possible association of CRISPR elements with potential virulence. Four types of CRISPR arrays were identified. CRISPR1 and CRISPR2 were present in all strains tested; 1 strain also had both CRISPR3 and CRISPR4, whereas 193 strains displayed a short, combined array, CRISPR3-4. A total of 3,353 spacers were identified, representing 528 distinct spacers. The average length of a spacer was 32 bp. Approximately one-half of the spacers (54%) were unique and found mostly in strains of less common serotypes. Overall, CRISPR spacer contents correlated well with STEC serotypes, and identical arrays were shared between strains with the same H type (O26:H11, O103:H11, and O111:H11). There was no association identified between the presence of subtype I-E cas and virulence genes, but the total number of spacers had a negative correlation with potential pathogenicity (P CRISPR-cas system and potential virulence needs to be determined on a broader scale, and the biological link will need to be established.

  14. Use of intradermal botulinum toxin to reduce sebum production and facial pore size.

    Science.gov (United States)

    Shah, Anil R

    2008-09-01

    Review the safety profile and subjective efficacy of intradermal botulinum toxin type A in facial pore size and sebum production. Retrospective analysis of 20 patients. Twenty consecutive patients with a single application of intradermal botulinum toxin type A were examined: Patients (17/20) noted an improvement in sebum production and a decrease in pores size at 1 month after injection. No complications were observed, and 17/20 patients were satisfied with the procedure. Preliminary data suggests that intradermal botulinum toxin may play a role in decreasing sebum production. Further quantitive study may be necessary to determine effects of intradermal botulinum toxin on pore size.

  15. A Review of Current Research and Knowledge Gaps in the Epidemiology of Shiga Toxin-Producing Escherichia coli and Salmonella spp. in Trinidad and Tobago

    Directory of Open Access Journals (Sweden)

    Anil K. Persad

    2018-04-01

    Full Text Available Salmonella and Shiga toxin-producing Escherichia coli are two of the main causes of foodborne disease globally, and while they have been implicated as possible causes of foodborne disease within the Caribbean region, the actual incidence is unknown. Trinidad and Tobago, one of the larger countries in the Caribbean, has an estimated annual foodborne disease burden of over 100,000 cases and, similar to other countries, the etiology of most of these cases is unknown. Both pathogens can reside as part of the normal gastrointestinal microflora of many wild and domestic animals, with animals acting as reservoirs, spillover hosts, or dead-end hosts. Carriage in animal species can be asymptomatic or, in the case of Salmonella in particular, there may be clinical manifestation in animals, which resemble the disease seen in humans. In this review, we will focus on the epidemiology of these two foodborne pathogens in Trinidad and Tobago and identify any knowledge gaps in the published literature. The filling of this critical knowledge void is essential for the development and implementation of appropriate mechanisms to reduce the dissemination and transmission of these pathogens, not only in Trinidad and Tobago, but also in the wider Caribbean.

  16. Shiga toxin-producing Escherichia coli in yaks (Bos grunniens from the Qinghai-Tibetan Plateau, China.

    Directory of Open Access Journals (Sweden)

    Xiangning Bai

    Full Text Available Shiga toxin (Stx-producing Escherichia coli (STEC are recognized as important human pathogens of public health concern. Many animals are the sources of STEC. In this study we determined the occurrence and characteristics of the STEC in yaks (Bos grunniens from the Qinghai-Tibetan plateau, China. A total of 728 yak fecal samples was collected from June to August, 2012 and was screened for the presence of the stx 1 and stx 2 genes by TaqMan real-time PCR after the sample was enriched in modified Tryptone Soya Broth. Of the 138 (18.96% stx 1 and/or stx 2-positive samples, 85 (61.59% were confirmed to have at least 1 STEC isolate present by culture isolation, from which 128 STEC isolates were recovered. All STEC isolates were serotyped, genotyped by pulsed-field gel electrophoresis (PFGE and characterized for the presence of 16 known virulence factors. Fifteen different O serogroups and 36 different O:H serotypes were identified in the 128 STEC isolates with 21 and 4 untypable for the O and H antigens respectively. One stx 1 subtype (stx 1a and 5 stx 2 subtypes (stx 2a, stx 2b, stx 2c, stx 2d and stx 2g were present in these STEC isolates. Apart from lpfA O157/OI-141, lpfA O157/OI-154, lpfA O113, katP and toxB which were all absent, other virulence factors screened (eaeA, iha, efa1, saa, paa, cnf1, cnf2, astA, subA, exhA and espP were variably present in the 128 STEC isolates. PFGE were successful for all except 5 isolates and separated them into 67 different PFGE patterns. For the 18 serotypes with 2 or more isolates, isolates of the same serotypes had the same or closely related PFGE patterns, demonstrating clonality of these serotypes. This study was the first report on occurrence and characteristics of STEC isolated from yaks (Bos grunniens from the Qinghai-Tibetan plateau, China, and extended the genetic diversity and reservoir host range of STEC.

  17. Characterization and zoonotic impact of Shiga toxin producing Escherichia coli in some wild bird species

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    Hanaa Mohamed Fadel

    2017-09-01

    Full Text Available Aim: Wild birds are considered silent vectors of some zoonotic water and food borne pathogens of public health significance. Owing to the importance of Shiga toxin producing Escherichia coli (STEC as the most pathogenic among the emerging diarrheagenic E. coli groups that can infect man; the present study was designed to detect the occurrence of STEC among wild birds in Egypt. Materials and Methods: A total of 177 intestinal content swab samples originating from five wild bird species were investigated for the presence of E. coli and STEC by standard culture methods. Suspect STEC isolates were further characterized by serotyping, random amplified polymorphic DNA polymerase chain reaction (RAPD PCR, antimicrobial resistance pattern and PCR detection of stx1, stx2, and eae genes. Results: A total of 30 suspect STEC isolates from 30 positive birds' samples were detected and identified on STEC CHROMagar (semi-captive pigeons, 15; house crows, 8; cattle egrets, 3; moorhens, 2; and house teals, 2. 25 isolates were grouped into 13 serogroups (O:20, O:25, O:26, O:27, O:63, O:78, O:111, O:114, O:125, O:128, O:142, O:153, and O:158, while five were rough strains. The distribution of STEC virulence genes among wild birds was as follows: 16 birds carried stx1 gene only (nine pigeons [28.1%], six crows [7.1%], and one cattle egret [5.6%]. stx1 and stx2 genes together were detected in four birds (one cattle egret [5.6%], two moorhens [6.1%], and one house teal, [10%]. Only one pigeon (3.1% possessed the three alleles. Disk diffusion test results showed that cefixime was the most effective against STEC serotypes with (93.3% sensitivity, followed by gentamycin (56.7%, and amoxicillin (50%. On the other hand, all the recovered STEC isolates were resistant to cefotaxime, doxycycline, cephalothin, and sulfisoxazole. RAPD fingerprinting using primers OPA-2 and OPA-9 showed that STEC isolates were heterogeneous; they yielded 30 and 27 different clusters

  18. Genetic makeup of Shiga toxin-producing Escherichia coli in relation to clinical symptoms and duration of shedding: a microarray analysis of isolates from Swedish children.

    Science.gov (United States)

    Matussek, A; Jernberg, C; Einemo, I-M; Monecke, S; Ehricht, R; Engelmann, I; Löfgren, S; Mernelius, S

    2017-08-01

    Shiga toxin (Stx)-producing Escherichia coli (STECs) cause non-bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome, and are the primary cause of acute renal failure in children worldwide. This study investigated the correlation of genetic makeup of STEC strains as revealed by DNA microarray to clinical symptoms and the duration of STEC shedding. All STEC isolated (n = 96) from patients <10 years of age in Jönköping County, Sweden from 2003 to 2015 were included. Isolates were characterized by DNA microarray, including almost 280 genes. Clinical data were collected through a questionnaire and by reviewing medical records. Of the 96 virulence genes (including stx) in the microarray, 62 genes were present in at least one isolate. Statistically significant differences in prevalence were observed for 21 genes when comparing patients with bloody diarrhea (BD) and with non-bloody stool (18 of 21 associated with BD). Most genes encode toxins (e.g., stx2 alleles, astA, toxB), adhesion factors (i.e. espB_O157, tir, eae), or secretion factors (e.g., espA, espF, espJ, etpD, nleA, nleB, nleC, tccP). Seven genes were associated with prolonged stx shedding; the presence of three genes (lpfA, senB, and stx1) and the absence of four genes (espB_O157, espF, astA, and intI1). We found STEC genes that might predict severe disease outcome already at diagnosis. This can be used to develop diagnostic tools for risk assessment of disease outcome. Furthermore, genes associated with the duration of stx shedding were detected, enabling a possible better prediction of length of STEC carriage after infection.

  19. Genotypes and phenotypes of Shiga toxin-producing Escherichia coli (STEC in Abeokuta, Southwestern Nigeria

    Directory of Open Access Journals (Sweden)

    Olowe OA

    2014-10-01

    Full Text Available Olugbenga Adekunle Olowe,1 Bukola W Aboderin,1,2 Olayinka O Idris,3 Victor O Mabayoje,4 Oluyinka O Opaleye,1 O Catherine Adekunle,1 Rita Ayanbolade Olowe,1 Paul Akinniyi Akinduti,5 Olusola Ojurongbe1 1Department of Medical Microbiology and Parasitology, College of Health Sciences, Osogbo, Osun State, Nigeria; 2Medical Microbiology Unit, Pathology Department, Federal Medical Centre, Abeokuta, Nigeria; 3Department of Microbiology, College of Sciences, Afe Babalola University, Ado Ekiti, Nigeria; 4Department of Haematology, College of Health Sciences, Ladoke Akintola University, Osogbo, Osun State, Nigeria; 5Department of Medical Microbiology and Parasitology, Olabisi Onabanjo University, Ago-Iwoye, Ogun State, Nigeria Purpose: To characterize the prevalence of hemolytic Shiga toxin-producing Escherichia coli (STEC with a multidrug-resistant pattern in different age groups in Abeokuta, Nigeria. Methods: Nonrepetitive E. coli isolates were collected from 202 subjects with or without evidence of diarrhea. Each isolate was biochemically identified and antimicrobial susceptibility testing was performed using the disk diffusion method. A sorbitol fermentation test of all the E. coli isolates was done and the minimum inhibitory concentration of suspected STEC was measured by the standard broth microdilution method to determine antibiotic resistance. The genotypes of stx1, stx2, and hlyA were determined by polymerase chain reaction assay. Results: The majority of subjects were aged ≥40 years (41.6% and were female (61.9%. Of the 202 subjects, 86.1% had STEC isolates (P<0.05. A high rate of STEC isolates resistant to amoxicillin (90.6%, cefotaxime (77.7%, and cefuroxime (75.7% was observed. Resistance to amoxicillin, gentamicin, and cefotaxime was demonstrated with a minimum inhibitory concentration >16 µg/mL in 13.9%, 11.4%, and 10.4% of the isolates, respectively. The prevalence of stx1, stx2, and hlyA was 13.9%, 6.9%, and 2.0%, respectively; 5.5% of

  20. Emerging types of Shiga toxin-producing E. coli (STEC O178 present in cattle, deer and humans from Argentina and Germany

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    Angelika eMiko

    2014-06-01

    Full Text Available More than 400 serotypes of Shiga toxin-producing Escherichia coli (STEC have been implicated in outbreaks and sporadic human diseases. In recent years STEC strains belonging to serogroup O178 have been commonly isolated from cattle and food of bovine origin in South America and Europe. In order to explore the significance of these STEC strains as potential human pathogens, 74 German and Argentinean E. coli O178 strains from animals, food and humans were characterized phenotypically and investigated for their serotypes, stx-genotypes and forty-three virulence-associated markers by a real-time PCR-microarray. The majority (n=66 of the O178 strains belonged to serotype O178:H19. The remaining strains divided into O178:H7 (n=6, O178:H10 (n=1 and O178:H16 (n=1. STEC O178:H19 strains were mainly isolated from cattle and food of bovine origin, but one strain was from a patient with hemolytic uremic syndrome (HUS. Genotyping of the STEC O178:H19 strains by pulsed-field gel electrophoresis revealed two major clusters of genetically highly related strains which differ in their stx-genotypes and non-Stx putative virulence traits, including adhesins, toxins and serine-proteases. Cluster A-strains including the HUS-strain (n=35 carried genes associated with severe disease in humans (stx2a, stx2d, ehxA, saa, subAB1, lpfAO113, terE combined with stx1a, espP, iha. Cluster B-strains (n=26 showed a limited repertoire of virulence genes (stx2c, pagC, lpfAO113, espP, iha. Among O178:H7 strains isolated from deer meat and patients with uncomplicated disease a new STEC variant was detected that is associated with the genotype stx1c/stx2b/ehxA/subAB2/espI/[terE]/espP/iha. None of the STEC O178 strains was positive for locus of enterocyte effacement (LEE- and nle-genes. Results indicate that STEC O178:H19 strains belong to the growing group of LEE-negative STEC that should be considered with respect to their potential to cause diseases in humans.

  1. Prolonged fecal shedding of Shiga toxin-producing Escherichia coli among children attending day-care centers in Argentina Excreción prolongada de Escherichia coli productor de toxina Shiga en niños que concurren a jardines maternales de Argentina

    Directory of Open Access Journals (Sweden)

    E. Miliwebsky

    2007-06-01

    Full Text Available In this report we describe the detection and duration of fecal shedding of Shiga toxin-producing Escherichia coli (STEC O157 and non-O157 in symptomatic and asymptomatic cases during four events occurred among children in day-care centers in Argentina. In each event, the cases were identified among children, family contacts and staff members of the Institution. The isolates were characterized by pheno-genotyping and subtyping methods. The STEC fecal shedding was prolonged and intermittent. Strains O157:H7 (1st event; O26:H11 (2nd event; O26:H11 (3rd event and O145:NM (4th event were shed during 23-30, 37, 31 and 19 days, respectively. Considering the possibility of STEC intermittent long-term shedding, symptomatic and asymptomatic individuals should be excluded from the Institution until two consecutive stool cultures obtained at least 48 h apart, test negative.En el presente trabajo se describe la detección y el tiempo de excreción de Escherichia coli productor de toxina Shiga (STEC O157 y no-O157 en casos sintomáticos y asintomáticos durante cuatro eventos ocurridos en jardines maternales de Argentina. En cada evento se identificaron los casos entre los niños, sus familiares y el personal del jardín. Los aislamientos fueron caracterizados por técnicas feno-genotípicas y de subtipificación. La excreción de STEC fue, en general, prolongada e intermitente. Cepas STEC O157:H7 (1er evento; O26:H11 (2do evento; O26:H11 (3er evento y O145:NM (4to evento fueron excretadas durante 23-30, 37, 31 y 19 días, respectivamente. Dadas las características de la excreción, no debe permitirse el reingreso a la institución de todo niño o adulto con infección por STEC, sintomático o asintomático, hasta no tener dos coprocultivos negativos sucesivos, con intervalos de 48 horas entre ellos.

  2. Development of a High Resolution Virulence Allelic Profiling (HReVAP) Approach Based on the Accessory Genome of Escherichia coli to Characterize Shiga-Toxin Producing E. coli (STEC)

    Science.gov (United States)

    Michelacci, Valeria; Orsini, Massimiliano; Knijn, Arnold; Delannoy, Sabine; Fach, Patrick; Caprioli, Alfredo; Morabito, Stefano

    2016-01-01

    Shiga-toxin producing Escherichia coli (STEC) strains possess a large accessory genome composed of virulence genes existing in multiple allelic variants, which sometimes segregate with specific STEC subpopulations. We analyzed the allelic variability of 91 virulence genes of STEC by Real Time PCR followed by melting curves analysis in 713 E. coli strains including 358 STEC. The 91 genes investigated were located on the locus of enterocyte effacement (LEE), OI-57, and OI-122 pathogenicity islands and displayed a total of 476 alleles in the study population. The combinations of the 91 alleles of each strain were termed allelic signatures and used to perform cluster analyses. We termed such an approach High Resolution Virulence Allelic Profiling (HReVAP) and used it to investigate the phylogeny of STEC of multiple serogroups. The dendrograms obtained identified groups of STEC segregating approximately with the serogroups and allowed the identification of subpopulations within the single groups. The study of the allelic signatures provided further evidence of the coevolution of the LEE and OI-122, reflecting the occurrence of their acquisition through a single event. The HReVAP analysis represents a sensitive tool for studying the evolution of LEE-positive STEC. PMID:26941726

  3. Effect of Cultured Celery Juice, Temperature, and Product Composition on the Inhibition of Proteolytic Clostridium botulinum Toxin Production.

    Science.gov (United States)

    Golden, Max C; Wanless, Brandon J; David, Jairus R D; Kottapalli, Bala; Lineback, D Scott; Talley, Ryan J; Glass, Kathleen A

    2017-08-01

    Clostridium botulinum may be of concern in prepared refrigerated meals, for which strict cold chain management cannot be guaranteed. This study evaluated the effect of temperature, product composition, and cultured celery juice powder (CCJP) as a source of nitrite on the inhibition of botulinum toxin formation in two experimental (meat- and vegetable-based) prepared meals. Data obtained from the challenge study were compared with a published mathematical model to determine whether the model is fail-safe with regard to the tested meals. Treatments were inoculated with proteolytic C. botulinum, vacuum packaged, cooked at 90°C for 10 min, and assayed for botulinum toxin at appropriate intervals in samples stored at 10, 15, or 20°C for up to 8 weeks. None of the treatments stored at 10°C for 8 weeks supported toxin production by proteolytic C. botulinum. The addition of CCJP delayed toxin production by 1 and 3 weeks in cauliflower potatoes and in Dijon pork, respectively, stored at 15°C. Toxin production was delayed by 1 week at 20°C when CCJP was added to the cauliflower potatoes. This study found that the predictive model was fail-safe but was overly conservative for the experimental meals described. Finally, this study confirms that product composition, the addition of nitrite via CCJP, storage time, and temperature play important roles in the inhibition of toxin formation by proteolytic C. botulinum.

  4. Toxin production in food as influenced by pH, thermal treatment and ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-06-03

    Jun 3, 2008 ... toxigenic properties and thirteen of the sixteen were positive for toxin production ... treatment and chemical preservatives on the growth rate and toxin ..... Clostridium botulinum in cooked pureed vegetables at Refrigerated.

  5. A polyclonal antibody based immunoassay detects seven subtypes of Shiga toxin 2 produced by Escherichia coli in human and environmental samples.

    Directory of Open Access Journals (Sweden)

    Xiaohua He

    Full Text Available BACKGROUND: Shiga toxin-producing Escherichia coli (STEC are frequent causes of severe human diseases ranging from diarrhea to hemolytic uremic syndrome. The existing strategy for detection of STEC relies on the unique sorbitol-negative fermentation property of the O157 strains, the most commonly identified serotype has been E. coli O157. It is becoming increasingly evident, however, that numerous non-O157 STEC serotypes also cause outbreaks and severe illnesses. It is necessary to have new methods that are capable of detecting all STEC strains. METHODS AND FINDINGS: Here we describe the development of a sandwich ELISA assay for detecting both O157 and non-O157 STECs by incorporating a novel polyclonal antibody (pAb against Stx2. The newly established immunoassay was capable of detecting Stx2a spiked in environmental samples with a limit of detection between 10 and 100 pg/mL in soil and between 100 and 500 pg/mL in feces. When applied to 36 bacterial strains isolated from human and environmental samples, this assay detected Stx2 in all strains that were confirmed to be stx2-positive by real-time PCR, demonstrating a 100% sensitivity and specificity. CONCLUSIONS: The sandwich ELISA developed in this study will enable any competent laboratory to identify and characterize Stx2-producing O157 and non-O157 strains in human and environmental samples, resulting in rapid diagnosis and patient care. The results of epitope mapping from this study will be useful for further development of a peptide-based antibody and vaccine.

  6. Antibody Response to Shiga Toxins in Argentinean Children with Enteropathic Hemolytic Uremic Syndrome at Acute and Long-Term Follow-Up Periods

    Science.gov (United States)

    Fernández-Brando, Romina J.; Bentancor, Leticia V.; Mejías, María Pilar; Ramos, María Victoria; Exeni, Andrea; Exeni, Claudia; Laso, María del Carmen; Exeni, Ramón; Isturiz, Martín A.; Palermo, Marina S.

    2011-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) infection is associated with a broad spectrum of clinical manifestations that include diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome (HUS). Systemic Stx toxemia is considered to be central to the genesis of HUS. Distinct methods have been used to evaluate anti-Stx response for immunodiagnostic or epidemiological analysis of HUS cases. The development of enzyme-linked immunosorbent assay (ELISA) and western blot (WB) assay to detect the presence of specific antibodies to Stx has introduced important advantages for serodiagnosis of HUS. However, application of these methods for seroepidemiological studies in Argentina has been limited. The aim of this work was to develop an ELISA to detect antibodies against the B subunit of Stx2, and a WB to evaluate antibodies against both subunits of Stx2 and Stx1, in order to analyze the pertinence and effectiveness of these techniques in the Argentinean population. We studied 72 normal healthy children (NHC) and 105 HUS patients of the urban pediatric population from the surrounding area of Buenos Aires city. Using the WB method we detected 67% of plasma from NHC reactive for Stx2, but only 8% for Stx1. These results are in agreement with the broad circulation of Stx2-expressing STEC in Argentina and the endemic behavior of HUS in this country. Moreover, the simultaneous evaluation by the two methods allowed us to differentiate acute HUS patients from NHC with a great specificity and accuracy, in order to confirm the HUS etiology when pathogenic bacteria were not isolated from stools. PMID:21559455

  7. Shiga toxin 1 induces on lipopolysaccharide-treated astrocytes the release of tumor necrosis factor-alpha that alter brain-like endothelium integrity.

    Directory of Open Access Journals (Sweden)

    Verónica I Landoni

    Full Text Available The hemolytic uremic syndrome (HUS is characterized by hemolytic anemia, thrombocytopenia and renal dysfunction. The typical form of HUS is generally associated with infections by Gram-negative Shiga toxin (Stx-producing Escherichia coli (STEC. Endothelial dysfunction induced by Stx is central, but bacterial lipopolysaccharide (LPS and neutrophils (PMN contribute to the pathophysiology. Although renal failure is characteristic of this syndrome, neurological complications occur in severe cases and is usually associated with death. Impaired blood-brain barrier (BBB is associated with damage to cerebral endothelial cells (ECs that comprise the BBB. Astrocytes (ASTs are inflammatory cells in the brain and determine the BBB function. ASTs are in close proximity to ECs, hence the study of the effects of Stx1 and LPS on ASTs, and the influence of their response on ECs is essential. We have previously demonstrated that Stx1 and LPS induced activation of rat ASTs and the release of inflammatory factors such as TNF-α, nitric oxide and chemokines. Here, we demonstrate that rat ASTs-derived factors alter permeability of ECs with brain properties (HUVECd; suggesting that functional properties of BBB could also be affected. Additionally, these factors activate HUVECd and render them into a proagregant state promoting PMN and platelets adhesion. Moreover, these effects were dependent on ASTs secreted-TNF-α. Stx1 and LPS-induced ASTs response could influence brain ECs integrity and BBB function once Stx and factors associated to the STEC infection reach the brain parenchyma and therefore contribute to the development of the neuropathology observed in HUS.

  8. Shiga Toxin 1 Induces on Lipopolysaccharide-Treated Astrocytes the Release of Tumor Necrosis Factor-alpha that Alter Brain-Like Endothelium Integrity

    Science.gov (United States)

    Landoni, Verónica I.; Schierloh, Pablo; de Campos Nebel, Marcelo; Fernández, Gabriela C.; Calatayud, Cecilia; Lapponi, María J.; Isturiz, Martín A.

    2012-01-01

    The hemolytic uremic syndrome (HUS) is characterized by hemolytic anemia, thrombocytopenia and renal dysfunction. The typical form of HUS is generally associated with infections by Gram-negative Shiga toxin (Stx)-producing Escherichia coli (STEC). Endothelial dysfunction induced by Stx is central, but bacterial lipopolysaccharide (LPS) and neutrophils (PMN) contribute to the pathophysiology. Although renal failure is characteristic of this syndrome, neurological complications occur in severe cases and is usually associated with death. Impaired blood-brain barrier (BBB) is associated with damage to cerebral endothelial cells (ECs) that comprise the BBB. Astrocytes (ASTs) are inflammatory cells in the brain and determine the BBB function. ASTs are in close proximity to ECs, hence the study of the effects of Stx1 and LPS on ASTs, and the influence of their response on ECs is essential. We have previously demonstrated that Stx1 and LPS induced activation of rat ASTs and the release of inflammatory factors such as TNF-α, nitric oxide and chemokines. Here, we demonstrate that rat ASTs-derived factors alter permeability of ECs with brain properties (HUVECd); suggesting that functional properties of BBB could also be affected. Additionally, these factors activate HUVECd and render them into a proagregant state promoting PMN and platelets adhesion. Moreover, these effects were dependent on ASTs secreted-TNF-α. Stx1 and LPS-induced ASTs response could influence brain ECs integrity and BBB function once Stx and factors associated to the STEC infection reach the brain parenchyma and therefore contribute to the development of the neuropathology observed in HUS. PMID:22479186

  9. Genetic Relatedness Among Shiga Toxin-Producing Escherichia coli Isolated Along the Animal Food Supply Chain and in Gastroenteritis Cases in Qatar Using Multilocus Sequence Typing.

    Science.gov (United States)

    Palanisamy, Srikanth; Chang, YuChen; Scaria, Joy; Penha Filho, Rafael Antonio Casarin; Peters, Kenlyn E; Doiphode, Sanjay H; Sultan, Ali; Mohammed, Hussni O

    2017-06-01

    Pathogenic Escherichia coli has been listed among the most important bacteria associated with foodborne illnesses around the world. We investigated the genetic relatedness among Shiga toxin-producing E. coli (STEC) isolated along the animal food supply chain and from humans diagnosed with gastroenteritis in Qatar. Samples were collected from different sources along the food supply chain and from patients admitted to the hospital with complaints of gastroenteritis. All samples were screened for the presence of E. coli O157:H7 and non-O157 STEC using a combination of bacterial enrichment and molecular detection techniques. A proportional sampling approach was used to select positive samples from each source for further multilocus sequence typing (MLST) analysis. Seven housekeeping genes described for STEC were amplified by polymerase chain reaction, sequenced, and analyzed by MLST. Isolates were characterized by allele composition, sequence type (ST) and assessed for epidemiologic relationship within and among different sources. Nei's genetic distance was calculated at the allele level between sample pools in each site downstream. E. coli O157:H7 occurred at a higher rate in slaughterhouse and retail samples than at the farm or in humans in our sampling. The ST171, an ST common to enterotoxigenic E. coli and atypical enteropathogenic E. coli, was the most common ST (15%) in the food supply chain. None of the genetic distances among the different sources was statistically significant. Enterohemorrhagic E. coli pathogenic strains are present along the supply chain at different levels and with varying relatedness. Clinical isolates were the most diverse, as expected, considering the polyclonal diversity in the human microbiota. The high occurrence of these food adulterants among the farm products suggests that implementation of sanitary measures at that level might reduce the risk of human exposure.

  10. Inhibitory effects of spices on growth and toxin production of toxigenic fungi.

    OpenAIRE

    Hitokoto, H; Morozumi, S; Wauke, T; Sakai, S; Kurata, H

    1980-01-01

    The inhibitory effects of 29 commercial powdered spices on the growth and toxin production of three species of toxigenic Aspergillus were observed by introducing these materials into culture media for mycotoxin production. Of the 29 samples tested, cloves, star anise seeds, and allspice completely inhibited the fungal growth, whereas most of the others inhibited only the toxin production. Eugenol extracted from cloves and thymol from thyme caused complete inhibition of the growth of both Aspe...

  11. A novel regulator controls Clostridium difficile sporulation, motility and toxin production.

    Science.gov (United States)

    Edwards, Adrianne N; Tamayo, Rita; McBride, Shonna M

    2016-06-01

    Clostridium difficile is an anaerobic pathogen that forms spores which promote survival in the environment and transmission to new hosts. The regulatory pathways by which C. difficile initiates spore formation are poorly understood. We identified two factors with limited similarity to the Rap sporulation proteins of other spore-forming bacteria. In this study, we show that disruption of the gene CD3668 reduces sporulation and increases toxin production and motility. This mutant was more virulent and exhibited increased toxin gene expression in the hamster model of infection. Based on these phenotypes, we have renamed this locus rstA, for regulator of sporulation and toxins. Our data demonstrate that RstA is a bifunctional protein that upregulates sporulation through an unidentified pathway and represses motility and toxin production by influencing sigD transcription. Conserved RstA orthologs are present in other pathogenic and industrial Clostridium species and may represent a key regulatory protein controlling clostridial sporulation. © 2016 John Wiley & Sons Ltd.

  12. Occurrence, genotyping, shiga toxin genes and associated risk factors of E. coli isolated from dairy farms, handlers and milk consumers.

    Science.gov (United States)

    Awadallah, M A; Ahmed, H A; Merwad, A M; Selim, M A

    2016-11-01

    The objectives of the current study were to determine the occurrence and genotypes of E. coli in dairy farms, workers and milk consumers and to evaluate risk factors associated with contamination of milk in dairy farms. Molecular characterization of shiga toxin associated genes and enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) finger printing of E. coli from different sources were also studied. Paired milk samples and rectal swabs from 125 dairy cows, rectal swabs from 82 calves and hand swabs from 45 dairy workers from five dairy farms were collected. In addition, 100 stool samples from 70 diarrheic and 30 healthy humans were collected and examined for the presence of E. coli. E. coli was isolated from milk (22.4%), dairy cattle feces (33.6%), calf feces (35.4%), dairy worker hand swabs (11.1%) and stools of milk consumers (2%, from diarrheic patients only). Only stx1 was identified in seven of 12 E. coli O125 isolated from different sources. High genetic diversity was determined (Simpson's index of diversity, D = 1) and E. coli O125 isolates were classified into 12 distinct profiles, E1-E12. The dendrogram analysis showed that two main clusters were generated. Mastitis in dairy cows was considered a risk factor associated with contamination of the produced milk with E. coli. The isolation of E. coli from rectal swabs of dairy cows and calves poses a zoonotic risk through consumption of unpasteurized contaminated dairy milk. Educational awareness should be developed to address risks related to consumption of raw milk. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Identification of genes expressed in cultures of E. coli lysogens carrying the Shiga toxin-encoding prophage Φ24B

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    Riley Laura M

    2012-03-01

    Full Text Available Abstract Background Shigatoxigenic E. coli are a global and emerging health concern. Shiga toxin, Stx, is encoded on the genome of temperate, lambdoid Stx phages. Genes essential for phage maintenance and replication are encoded on approximately 50% of the genome, while most of the remaining genes are of unknown function nor is it known if these annotated hypothetical genes are even expressed. It is hypothesized that many of the latter have been maintained due to positive selection pressure, and that some, expressed in the lysogen host, have a role in pathogenicity. This study used Change Mediated Antigen Technology (CMAT™ and 2D-PAGE, in combination with RT-qPCR, to identify Stx phage genes that are expressed in E. coli during the lysogenic cycle. Results Lysogen cultures propagated for 5-6 hours produced a high cell density with a low proportion of spontaneous prophage induction events. The expression of 26 phage genes was detected in these cultures by differential 2D-PAGE of expressed proteins and CMAT. Detailed analyses of 10 of these genes revealed that three were unequivocally expressed in the lysogen, two expressed from a known lysogenic cycle promoter and one uncoupled from the phage regulatory network. Conclusion Propagation of a lysogen culture in which no cells at all are undergoing spontaneous lysis is impossible. To overcome this, RT-qPCR was used to determine gene expression profiles associated with the growth phase of lysogens. This enabled the definitive identification of three lambdoid Stx phage genes that are expressed in the lysogen and seven that are expressed during lysis. Conservation of these genes in this phage genome, and other Stx phages where they have been identified as present, indicates their importance in the phage/lysogen life cycle, with possible implications for the biology and pathogenicity of the bacterial host.

  14. Multiplex real-time PCR assay for detection of Escherichia coli O157:H7 and screening for non-O157 Shiga toxin-producing E. coli.

    Science.gov (United States)

    Li, Baoguang; Liu, Huanli; Wang, Weimin

    2017-11-09

    Shiga toxin-producing Escherichia coli (STEC), including E. coli O157:H7, are responsible for numerous foodborne outbreaks annually worldwide. E. coli O157:H7, as well as pathogenic non-O157:H7 STECs, can cause life-threating complications, such as bloody diarrhea (hemolytic colitis) and hemolytic-uremic syndrome (HUS). Previously, we developed a real-time PCR assay to detect E. coli O157:H7 in foods by targeting a unique putative fimbriae protein Z3276. To extend the detection spectrum of the assay, we report a multiplex real-time PCR assay to specifically detect E. coli O157:H7 and screen for non-O157 STEC by targeting Z3276 and Shiga toxin genes (stx1 and stx2). Also, an internal amplification control (IAC) was incorporated into the assay to monitor the amplification efficiency. The multiplex real-time PCR assay was developed using the Life Technology ABI 7500 System platform and the standard chemistry. The optimal amplification mixture of the assay contains 12.5 μl of 2 × Universal Master Mix (Life Technology), 200 nM forward and reverse primers, appropriate concentrations of four probes [(Z3276 (80 nM), stx1 (80 nM), stx2 (20 nM), and IAC (40 nM)], 2 μl of template DNA, and water (to make up to 25 μl in total volume). The amplification conditions of the assay were set as follows: activation of TaqMan at 95 °C for 10 min, then 40 cycles of denaturation at 95 °C for 10 s and annealing/extension at 60 °C for 60 s. The multiplex assay was optimized for amplification conditions. The limit of detection (LOD) for the multiplex assay was determined to be 200 fg of bacterial DNA, which is equivalent to 40 CFU per reaction which is similar to the LOD generated in single targeted PCRs. Inclusivity and exclusivity determinants were performed with 196 bacterial strains. All E. coli O157:H7 (n = 135) were detected as positive and all STEC strains (n = 33) were positive for stx1, or stx2, or stx1 and stx2 (Table 1). No cross reactivity was detected with Salmonella

  15. Isolation, Characterization and Antibiotic Resistance of Shiga Toxin-Producing Escherichia coli in Hamburger and Evolution of Virulence Genes stx1, stx2, eaeA and hly by Multiplex PCR

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    Mohammad Kargar

    2013-09-01

    Full Text Available Background & Objectives: Shiga toxin-producing Escherichia coli (STEC O157:H7 have emerged as pathogens that can cause food-borne infections and severe and potentially fatal illnesses in humans. E.coli O157:H7 colonizes the digestive tract of cattle and is transmitted to humans by food and water. The objectives of this study were to characterize the prevalence of E.coli O157:H7 isolates in hamburger in Shiraz and to test their antimicrobial sensitivity. Material & Methods: In this research, 428 samples of hamburger were collected from 7 main factories of meat products and enriched in TSB with novobiocin medium at 37ºC. Fermentation of sorbitol and lactose and activities of β- glucuronidase of separated bacteria were examined by using the SMAC and VRBA media and CHROMagar medium. Then isolation of E.coli O157:H7 was confirmed with the use of specific antisera; and with the multiplex PCR method, the presence of E.coli O157:H7 virulence genes – including stx1, stx2, eaeA, and hly – was analyzed. Finally, antibiotic resistance strains were tested with disk diffusion methods. Results: Out of all the examined samples, 264 (61.68% sorbitol-negative bacteria were separated in the CT-SMAC medium. After evaluation with specific antisera, the rate of the recognition of E.coli O157:H7 was 5 (1.17%. The stx1 and eaeA genes were diagnosed in 2 (0.47% cases of these samples. All the isolated bacteria were resistant to penicillin, clindamycin, and erythromycin antibiotics.Conclusion: The presence of STEC in animal products suggests that they may be a potential hazard for human health. A regular monitoring of STEC O157, mainly in hamburger, should be performed to prevent a possible consumer health threat.

  16. Prevalence and characteristics of Shiga toxin-producing Escherichia coli in finishing pigs: Implications on public health.

    Science.gov (United States)

    Cha, Wonhee; Fratamico, Pina M; Ruth, Leah E; Bowman, Andrew S; Nolting, Jacqueline M; Manning, Shannon D; Funk, Julie A

    2018-01-02

    Shiga toxin-producing Escherichia coli (STEC) are important food-borne pathogens, which can cause serious illnesses, including hemorrhagic colitis and hemolytic uremic syndrome. To study the epidemiology of STEC in finishing pigs and examine the potential risks they pose for human STEC infections, we conducted a longitudinal cohort study in three finishing sites. Six cohorts of pigs (2 cohorts/site, 20 pigs/cohort) were randomly selected, and fecal samples (n=898) were collected every two weeks through their finishing period. Eighty-two pigs (68.3%) shed STEC at least once, and the proportion of STEC-positive pigs varied across sites (50-97.5%) and cohorts (15-100%). Clinically important serotypes, O157:H7 (stx 2c , eae) and O26:H11 (stx 1a , eae), were recovered from two pigs at sites C and A, respectively. The most common serotype isolated was O59:H21 (stx 2e ), which was particularly prevalent in site B as it was recovered from all STEC positive pigs (n=39). Each cohort showed different patterns of STEC shedding, which were associated with the prevalent serotype. The median shedding duration of STEC in pigs was 28days, consistent with our prior study. However, among pigs shedding O59:H21 at least once, pigs in cohort B2 had a significantly longer shedding duration of 42days (P<0.05) compared to other cohorts. Stx2e was the most commonly observed stx variant in finishing pigs (93.9%), in accordance with the previous studies. Stx2e has been reported to be significantly associated with edema disease in pigs, however, the pathogenicity in humans warrants further investigations. Nonetheless, our findings affirm that pigs are an important reservoir for human STEC infections, and that the circulating serotypes in a cohort and site management factors may significantly affect the prevalence of STEC. Molecular characterization of STEC isolates and epidemiological studies to identify risk factors for shedding in pigs are strongly warranted to further address the

  17. Toxin production ability of Bacillus cereus strains from food product of Ukraine

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    I. Pylypenko

    2017-10-01

    Full Text Available Potential pathogens of foodborne toxic infections – bacterial contaminants Bacillus cereus isolated from plant raw materials and food products from the Ukrainian region were investigated. When determining of the proportion of isolated bacilli from the plant samples, it was established that the epidemiologically significant microorganisms of Bacillus cereus as agents of food poisoning are the second largest. The average value of contaminated samples of Ukrainian plant raw materials and processed products with Bacillus cereus is 36,2 %. The ability of Bacillus cereus strains identified by a complex of morphological, tinctorial, cultural and biochemical properties, to produce specific emetic and enterotoxins was studied. Molecular genetic diagnosis and detection of the toxin-producing ability of isolated 42 Bacillus cereus strains showed both the possibility of their rapid identification and the presence of specific toxicity genes. Multiplex polymerase chain reaction (PCR was carried out with specific primers to detect toxicity determined of various bacilli genes: nheA, hblD, cytK, cesВ. The distribution of toxigenic genes is significantly different among the Bacillus cereus isolates from various sources. The nheA, hblD and cytK enterotoxin genes were detected in 100, 83,3 and 61,9 % of the investigated strains of Bacillus cereus, respectively. The cesB gene encoding emetic toxin was detected in 4,8 % of  strains. Molecular-genetic PCR-method confirmed that all the isolated strains belong to the Bacillus cereus group, and the ability to produce toxins can be attributed to five groups. The main toxins that produce the investigated Bacillus cereus strains were nhe and hbl enterotoxins encoded by the corresponding genes of nheA and hblD. The enterotoxic type of Bacillus cereus was predominant in Ukrainian region.  Studies of domestic plant food raw materials and products have confirmed the need to improve microbiological control of product safety

  18. Detección y caracterización de Escherichia coli productor de toxina Shiga a partir de casos clínicos y de alimentos en Uruguay Detection and characterization of Shiga toxin - producing Escherichia coli from clinical cases and food in Uruguay

    Directory of Open Access Journals (Sweden)

    G. Varela

    2008-06-01

    Full Text Available Establecimos la frecuencia de aislamiento de Escherichia coli productor de toxina Shiga (STEC a partir de muestras clínicas y de alimentos, así como las características fenotípicas y genotípicas de las cepas recuperadas. Se analizaron 198 muestras fecales de niños con diarrea sanguinolenta (DS, 14 muestras fecales de niños con síndrome urémico hemolítico (SUH y 220 muestras de carne picada. También se estudiaron 4 cepas STEC aisladas de alimentos embutidos. Se recuperó STEC de 3 (1,5% de los niños con DS, de 1 (7% niño con SUH y de 4 (1,8% de las muestras de carne picada. Todas las cepas fueron eae y ehxA positivas. Los serotipos detectados fueron: O157:H7 (9 cepas, O26:H11 (2 cepas, O111:NM (1 cepa y O145:HNT (1 cepa. Todas las cepas O157:H7 portaron el subtipo eae-g1; las cepas O26:H11 y O145:HNT portaron el subtipo eae-b1 y la cepa O111:NM portó el subtipo eae-g2/q. Las cepas STEC del mismo serogrupo mostraron alta diversidad genética. En Uruguay STEC no sería agente frecuente de diarrea con sangre en niños. Sin embargo, las cepas recuperadas presentaron los genes asociados con enfermedad severa y 2 de los 3 niños infectados con STEC evolucionaron a SUH. La carne picada y otros alimentos serían vehículos importantes de O157:H7.We have assessed the frequency of Shiga toxin-producing Escherichia coli (STEC in clinical and food samples as well as studied the genotypic and phenotypic characteristics of the recovered strains. One hundred ninety eight fecal samples from children with bloody diarrhea (BD, 14 from children with hemolytic uremic syndrome (HUS, 220 ground beef samples and 4 STEC isolates from other beef-derived products were analyzed. The STEC strains were isolated from 3 (1.5% children with bloody diarrhea, 1 (7% from a child with HUS and 4 (1.8% from ground beef samples. All strains were eae and ehxA positive. The serotypes found were: O157:H7 (9 strains, O26:H11 (2, O111: NM (1 and O145:HNT (1. All O157:H7 STEC

  19. Epidemiological analysis of a cluster within the outbreak of Shiga toxin-producing Escherichia coli serotype O104:H4 in Northern Germany, 2011.

    Science.gov (United States)

    Scharlach, Martina; Diercke, Michaela; Dreesman, Johannes; Jahn, Nicola; Krieck, Manuela; Beyrer, Konrad; Claußen, Katja; Pulz, Matthias; Floride, Regina

    2013-06-01

    In May 2011 one of the worldwide largest outbreaks of haemolytic uraemic syndrome (HUS) and bloody diarrhoea caused by Shiga toxin-producing Escherichia coli (STEC) serotype O104:H4 occurred in Germany. One of the most affected federal states was Lower Saxony. We present the investigation of a cluster of STEC and HUS cases within this outbreak by means of a retrospective cohort study. After a 70th birthday celebration which took place on 7th of May 2011 among 72 attendants seven confirmed cases and four probable cases were identified, two of them developed HUS. Median incubation period was 10 days. Only 35 persons (48.6%) definitely answered the question whether they had eaten the sprouts that were used for garnishing the salad. Univariable analysis revealed different food items, depending on the case definition, with Odds Ratio (OR)>1 indicating an association with STEC infection, but multivariable logistic regression showed no increased risk for STEC infection for any food item and any case definition. Sprouts as the source for the infection had to be assumed based on the results of a tracing back of the delivery ways from the catering company to the sprouts producer who was finally identified as the source of the entire German outbreak. In this large outbreak several case-control studies failed to identify the source of infection. Copyright © 2012 Elsevier GmbH. All rights reserved.

  20. Toxin production in food as influenced by pH, thermal treatment and ...

    African Journals Online (AJOL)

    Toxin production in food as influenced by pH, thermal treatment and chemical ... carrot, spinach, pepper, tomato, onion and cooked food samples (rice, yam, beans, ... on the growth rate and toxin elaboration of E. coli, K. aerogenes, C. freundii, ...

  1. Mouldy feed, mycotoxins and Shiga toxin - producing Escherichia coli colonization associated with Jejunal Hemorrhage Syndrome in beef cattle

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    Masson Luke

    2011-06-01

    Full Text Available Abstract Background Both O157 and non-O157 Shiga toxin - producing Escherichia coli (STECs cause serious human disease outbreaks through the consumption of contaminated foods. Cattle are considered the main reservoir but it is unclear how STECs affect mature animals. Neonatal calves are the susceptible age class for STEC infections causing severe enteritis. In an earlier study, we determined that mycotoxins and STECs were part of the disease complex for dairy cattle with Jejunal Hemorrhage Syndrome (JHS. For STECs to play a role in the development of JHS, we hypothesized that STEC colonization should also be evident in beef cattle with JHS. Aggressive medical and surgical therapies are effective for JHS, but rely on early recognition of clinical signs for optimal outcomes suggesting that novel approaches must be developed for managing this disease. The main objective of this study was to confirm that mouldy feeds, mycotoxins and STEC colonization were associated with the development of JHS in beef cattle. Results Beef cattle developed JHS after consuming feed containing several types of mycotoxigenic fungi including Fusarium poae, F. verticillioides, F. sporotrichioides, Penicillium roqueforti and Aspergillus fumigatus. Mixtures of STECs colonized the mucosa in the hemorrhaged tissues of the cattle and no other pathogen was identified. The STECs expressed Stx1 and Stx2, but more significantly, Stxs were also present in the blood collected from the lumen of the hemorrhaged jejunum. Feed extracts containing mycotoxins were toxic to enterocytes and 0.1% of a prebiotic, Celmanax Trademark, removed the cytotoxicity in vitro. The inclusion of a prebiotic in the care program for symptomatic beef calves was associated with 69% recovery. Conclusions The current study confirmed that STECs and mycotoxins are part of the disease complex for JHS in beef cattle. Mycotoxigenic fungi are only relevant in that they produce the mycotoxins deposited in the feed. A

  2. Comparative Genomics of Recent Shiga Toxin-Producing Escherichia coli O104:H4: Short-Term Evolution of an Emerging Pathogen

    Science.gov (United States)

    Grad, Yonatan H.; Godfrey, Paul; Cerquiera, Gustavo C.; Mariani-Kurkdjian, Patricia; Gouali, Malika; Bingen, Edouard; Shea, Terrence P.; Haas, Brian J.; Griggs, Allison; Young, Sarah; Zeng, Qiandong; Lipsitch, Marc; Waldor, Matthew K.; Weill, François-Xavier; Wortman, Jennifer R.; Hanage, William P.

    2013-01-01

    ABSTRACT The large outbreak of diarrhea and hemolytic uremic syndrome (HUS) caused by Shiga toxin-producing Escherichia coli O104:H4 in Europe from May to July 2011 highlighted the potential of a rarely identified E. coli serogroup to cause severe disease. Prior to the outbreak, there were very few reports of disease caused by this pathogen and thus little known of its diversity and evolution. The identification of cases of HUS caused by E. coli O104:H4 in France and Turkey after the outbreak and with no clear epidemiological links raises questions about whether these sporadic cases are derived from the outbreak. Here, we report genome sequences of five independent isolates from these cases and results of a comparative analysis with historical and 2011 outbreak isolates. These analyses revealed that the five isolates are not derived from the outbreak strain; however, they are more closely related to the outbreak strain and each other than to isolates identified prior to the 2011 outbreak. Over the short time scale represented by these closely related organisms, the majority of genome variation is found within their mobile genetic elements: none of the nine O104:H4 isolates compared here contain the same set of plasmids, and their prophages and genomic islands also differ. Moreover, the presence of closely related HUS-associated E. coli O104:H4 isolates supports the contention that fully virulent O104:H4 isolates are widespread and emphasizes the possibility of future food-borne E. coli O104:H4 outbreaks. PMID:23341549

  3. Efecto citotóxico de la toxina shiga tipo 2 y su subunidad b en células epiteliales tubulares renales humanas en cultivo Cytotoxic effect of Shiga toxin type 2 and its B subunit on human renal tubular epithelial cell cultures

    Directory of Open Access Journals (Sweden)

    Virginia Pistone Creydt

    2005-04-01

    Full Text Available Escherichia coli enterohemorrágica productora de toxina Shiga (Stx causa diarrea acuosa, colitis hemorrágica y síndrome urémico hemolítico (SUH. En Argentina, el SUH es la principal causa de insuficiencia renal en niños. El objetivo de este trabajo fue estudiar la toxicidad de Stx tipo 2 (Stx2 y su subunidad B (Stx2B en células epiteliales tubulares renales humanas (CERH, en presencia y ausencia de factores inflamatorios. Los efectos citotóxicos se evaluaron como alteración de la funcionalidad del epitelio; daños histológicos; viabilidad celular; síntesis de proteínas y apoptosis celular. Los resultados muestran que Stx2 regula el pasaje de agua a través de CERH a tiempos menores de 1h de incubación. A tiempos mayores, hasta 72 hs, el estudio de la morfología, la viabilidad, la síntesis de proteínas y la apoptosis demostró que las CERH fueron sensibles a la acción citotóxica de Stx2 y Stx2B de una manera dosis y tiempo dependiente. Estos efectos fueron potenciados por lipopolisacáridos bacterianos (LPS, IL-1b, y butirato.Shiga toxin (Stx-producing E.coli causing watery diarrhea, hemorrhagic colitis and hemolytic-uremic syndrome (HUS. In Argentina, HUS is the most common cause of acute renal failure in children. The purpose of the present study was to examine the cytotoxicity of Stx type 2 (Stx2 and its B subunit (Stx2B on human renal tubular epithelial cells (HRTEC, in the presence and absence of inflammatory factors. Cytotoxic effects were assessed in terms of functionality of the epithelium, histological damage, cell viability, protein synthesis and cellular apoptosis. Results show that Stx2 regulates the passage of water through the HRTEC within an incubation period of 1h. Within longer periods, up to 72 hours, the study of morphology, viability, protein synthesis and apoptosis shows that HRTEC were sensitive to the cytotoxic action of Stx2 and Stx2B in a dose- and time-dependent manner. These effects were potentiated by

  4. Occurrence and characterization of Shiga toxin-producing Escherichia coli in raw meat, raw milk, and street vended juices in Bangladesh.

    Science.gov (United States)

    Islam, Mohammad A; Mondol, Abdus S; Azmi, Ishrat J; de Boer, Enne; Beumer, Rijkelt R; Zwietering, Marcel H; Heuvelink, Annet E; Talukder, Kaisar A

    2010-11-01

    The major objective of this study was to investigate the prevalence of Shiga toxin (Stx)-producing Escherichia coli (STEC) in different types of food samples and to compare their genetic relatedness with STEC strains previously isolated from animal sources in Bangladesh. We investigated a total of 213 food samples, including 90 raw meat samples collected from retail butcher shops, 20 raw milk samples from domestic cattle, and 103 fresh juice samples from street vendors in Dhaka city. We found that more than 68% (n = 62) of the raw meat samples were positive for the stx gene(s); 34% (n = 21) of buffalo meats and 66% (n = 41) of beef. Approximately 10% (n = 2) of the raw milk and 8% (n = 8) of the fresh juice samples were positive for stx. We isolated STEC O157 from seven meat samples (7.8%), of which two were from buffalo meats and five from beef; and no other STEC serotypes could be isolated. We could not isolate STEC from any of the stx-positive raw milk and juice samples. The STEC O157 isolates from raw meats were positive for the stx(2), eae, katP, etpD, and enterohemorrhagic E. coli hly virulence genes, and they belonged to three different phage types: 8 (14.3%), 31 (42.8%), and 32 (42.8%). Pulsed-field gel electrophoresis (PFGE) typing revealed six distinct patterns among seven isolates of STEC O157, suggesting a heterogeneous clonal diversity. Of the six PFGE patterns, one was identical and the other two were ≥90% related to PFGE patterns of STEC O157 strains previously isolated from animal feces, indicating that raw meats are readily contaminated with fecal materials. This study represents the first survey of STEC in the food chain in Bangladesh.

  5. Standardization of process for increased production of pure and potent tetanus toxin

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    Chellamani Muniandi

    2013-09-01

    Full Text Available When stationary pot culture was replaced by submerged cultivation of Clostridium tetani, an anaerobic organism, in afermentor using a vibromixer and optimum supply of sterile air to the headspace of the fermentor to flush out the accumulatedgases, a significant increase in the tetanus toxin yield in a short time cultivation (about 5 to 6 days against8 days was noticed. It was found that under optimal conditions of temperature, vibromixing, surface aeration, and analkaline pH favored toxin release. Furthermore, to enhance the production volume, fermentor culture is more suitable.The tetanus toxin was produced with good Limes flocculation (Lf titre and high antigenic purity. Under optimal conditions,the papain digest broth was successfully substituted in place of N.Z Case for the production of pure and potenttetanus toxin. J Microbiol Infect Dis 2013; 3(3: 133-139Key words: Clostridium tetani, modified mueller miller medium, papain digest, limes flocculation

  6. Effect of Bacillus thuringiensis parasporal toxin on stimulating of IL-2 and IL-5 cytokines production

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    Marzieh Soleimany

    2018-03-01

    Full Text Available Introduction:Bacillus thuringiensis, is a Gram-positive spore-forming bacterium that produces crystalline parasporal protein (Cry during sporulation. Some of these Cry toxins do not show cytotoxicity against insects but they are capable to kill some human and animal cancer cells. The aim of this study was to verify whether cytocidal parasporal of B thuringiensis strains have immunostimulatory activity on human peripheral blood mononuclear cells (PBMNC and to evaluate the ability of IL-2 and IL-5 production. Materials and methods: B. thuringiensis toxin with cytocidal activity was isolated and treated with proteinase K. PBMNC was cultured and treated with activated crystal proteins. We evaluated the ability of different cytokines production with Flow Cytometry. Results: In this study, immune stimulatory toxins Cry1 were distinguished. This toxin can stimulate production of cytokines IL-2 and stop production of IL-5. Discussion and conclusion: According to anti-cancer effect of B. thuringiensis toxins and also immune stimulatory effect, with more research these toxins can be introduced as immunotherapy drug in cancer treatment.

  7. Effect of headspace CO2 concentration on toxin production by Clostridium botulinum in MAP, irradiated fresh pork

    International Nuclear Information System (INIS)

    Lambert, A.D.; Smith, J.P.; Dodds, K.L.

    1991-01-01

    The effects of five initial levels of CO2 (15, 30, 45, 60, and 75%) and three irradiation doses (0, 0.5, and 1.0 kGy) on toxin production by Clostridium botulinum in inoculated fresh pork were studied using factorial design experiments. Headspace CO2 levels increased in all samples during storage at 15 degrees C. In most treatments, spoilage preceded toxigenesis. Toxin production occurred faster in samples initially packaged with 15 to 30% of CO2 while higher levels of CO2 (45-75%) delayed toxin production. Low-dose irradiation delayed toxin production at all levels of CO2 in the package headspace. Contrary to expectations, including a CO2 absorbent in the package enhanced toxin production by C. botulinum. This was attributed to production of H2 by the CO2 absorbent, possibly resulting in a decrease in the oxido-reduction potential of the meat

  8. Development of an ELISA microarray assay for the sensitive and simultaneous detection of ten biodefense toxins.

    Energy Technology Data Exchange (ETDEWEB)

    Jenko, Kathryn; Zhang, Yanfeng; Kostenko, Yulia; Fan, Yongfeng; Garcia-Rodriguez, Consuelo; Lou, Jianlong; Marks, James D.; Varnum, Susan M.

    2014-10-21

    Plant and microbial toxins are considered bioterrorism threat agents because of their extreme toxicity and/or ease of availability. Additionally, some of these toxins are increasingly responsible for accidental food poisonings. The current study utilized an ELISA-based protein antibody microarray for the multiplexed detection of ten biothreat toxins, botulinum neurotoxins (BoNT) A, B, C, D, E, F, ricin, shiga toxins 1 and 2 (Stx), and staphylococcus enterotoxin B (SEB), in buffer and complex biological matrices. The multiplexed assay displayed a sensitivity of 1.3 pg/mL (BoNT/A, BoNT/B, SEB, Stx-1 and Stx-2), 3.3 pg/mL (BoNT/C, BoNT/E, BoNT/F) and 8.2 pg/mL (BoNT/D, ricin). All assays demonstrated high accuracy (75-120 percent recovery) and reproducibility (most coefficients of variation < 20%). Quantification curves for the ten toxins were also evaluated in clinical samples (serum, plasma, nasal fluid, saliva, stool, and urine) and environmental samples (apple juice, milk and baby food) with overall minimal matrix effects. The multiplex assays were highly specific, with little crossreactivity observed between the selected toxin antibodies. The results demonstrate a multiplex microarray that improves current immunoassay sensitivity for biological warfare agents in buffer, clinical, and environmental samples.

  9. Detection and Characterization of Shiga Toxin Producing Escherichia coli, Salmonella spp., and Yersinia Strains from Human, Animal, and Food Samples in San Luis, Argentina

    Science.gov (United States)

    Favier, Gabriela Isabel; Lucero Estrada, Cecilia; Cortiñas, Teresa Inés; Escudero, María Esther

    2014-01-01

    Shiga toxin producing Escherichia coli (STEC), Salmonella spp., and Yersinia species was investigated in humans, animals, and foods in San Luis, Argentina. A total of 453 samples were analyzed by culture and PCR. The antimicrobial susceptibility of all the strains was studied, the genomic relationships among isolates of the same species were determined by PFGE, and the potencial virulence of Y. enterocolitica strains was analyzed. Yersinia species showed higher prevalence (9/453, 2.0%, 95% CI, 0.7–3.3%) than STEC (4/453, 0.9%, 95% CI, 0–1.8%) and Salmonella spp. (3/453, 0.7%, 95% CI, 0–1.5%). Y. enterocolitica and Y. intermedia were isolated from chicken carcasses (6/80, 7.5%, 95% CI, 1.5–13.5%) and porcine skin and bones (3/10, 30%, 95% CI, 0–65%). One STEC strain was recovered from human feces (1/70, 1.4%, 95% CI, 0–4.2%) and STEC stx1/stx2 genes were detected in bovine stools (3/129, 2.3%, 95% CI, 0–5.0%). S. Typhimurium was isolated from human feces (1/70, 1.4%, 95% CI, 0–4.2%) while one S. Newport and two S. Gaminara strains were recovered from one wild boar (1/3, 33%, 95% CI, 0–99%). The knowledge of prevalence and characteristics of these enteropathogens in our region would allow public health services to take adequate preventive measures. PMID:25177351

  10. Review of the inhibition of biological activities of food-related selected toxins by natural compounds.

    Science.gov (United States)

    Friedman, Mendel; Rasooly, Reuven

    2013-04-23

    There is a need to develop food-compatible conditions to alter the structures of fungal, bacterial, and plant toxins, thus transforming toxins to nontoxic molecules. The term 'chemical genetics' has been used to describe this approach. This overview attempts to survey and consolidate the widely scattered literature on the inhibition by natural compounds and plant extracts of the biological (toxicological) activity of the following food-related toxins: aflatoxin B1, fumonisins, and ochratoxin A produced by fungi; cholera toxin produced by Vibrio cholerae bacteria; Shiga toxins produced by E. coli bacteria; staphylococcal enterotoxins produced by Staphylococcus aureus bacteria; ricin produced by seeds of the castor plant Ricinus communis; and the glycoalkaloid α-chaconine synthesized in potato tubers and leaves. The reduction of biological activity has been achieved by one or more of the following approaches: inhibition of the release of the toxin into the environment, especially food; an alteration of the structural integrity of the toxin molecules; changes in the optimum microenvironment, especially pH, for toxin activity; and protection against adverse effects of the toxins in cells, animals, and humans (chemoprevention). The results show that food-compatible and safe compounds with anti-toxin properties can be used to reduce the toxic potential of these toxins. Practical applications and research needs are suggested that may further facilitate reducing the toxic burden of the diet. Researchers are challenged to (a) apply the available methods without adversely affecting the nutritional quality, safety, and sensory attributes of animal feed and human food and (b) educate food producers and processors and the public about available approaches to mitigating the undesirable effects of natural toxins that may present in the diet.

  11. Phenotypic H-Antigen Typing by Mass Spectrometry Combined with Genetic Typing of H Antigens, O Antigens, and Toxins by Whole-Genome Sequencing Enhances Identification of Escherichia coli Isolates.

    Science.gov (United States)

    Cheng, Keding; Chui, Huixia; Domish, Larissa; Sloan, Angela; Hernandez, Drexler; McCorrister, Stuart; Robinson, Alyssia; Walker, Matthew; Peterson, Lorea A M; Majcher, Miles; Ratnam, Sam; Haldane, David J M; Bekal, Sadjia; Wylie, John; Chui, Linda; Tyler, Shaun; Xu, Bianli; Reimer, Aleisha; Nadon, Celine; Knox, J David; Wang, Gehua

    2016-08-01

    Mass spectrometry-based phenotypic H-antigen typing (MS-H) combined with whole-genome-sequencing-based genetic identification of H antigens, O antigens, and toxins (WGS-HOT) was used to type 60 clinical Escherichia coli isolates, 43 of which were previously identified as nonmotile, H type undetermined, or O rough by serotyping or having shown discordant MS-H and serotyping results. Whole-genome sequencing confirmed that MS-H was able to provide more accurate data regarding H antigen expression than serotyping. Further, enhanced and more confident O antigen identification resulted from gene cluster based typing in combination with conventional typing based on the gene pair comprising wzx and wzy and that comprising wzm and wzt The O antigen was identified in 94.6% of the isolates when the two genetic O typing approaches (gene pair and gene cluster) were used in conjunction, in comparison to 78.6% when the gene pair database was used alone. In addition, 98.2% of the isolates showed the existence of genes for various toxins and/or virulence factors, among which verotoxins (Shiga toxin 1 and/or Shiga toxin 2) were 100% concordant with conventional PCR based testing results. With more applications of mass spectrometry and whole-genome sequencing in clinical microbiology laboratories, this combined phenotypic and genetic typing platform (MS-H plus WGS-HOT) should be ideal for pathogenic E. coli typing. Copyright © 2016 Cheng et al.

  12. Detection of Shiga toxin-producing Escherichia coli (STEC) O157:H7, O26, O45, O103, O111, O121, and O145, and Salmonella in retail raw ground beef using the DuPont™ BAX® system.

    Science.gov (United States)

    Wasilenko, Jamie L; Fratamico, Pina M; Sommers, Christopher; DeMarco, Daniel R; Varkey, Stephen; Rhoden, Kyle; Tice, George

    2014-01-01

    Shiga toxin-producing Escherichia coli (STEC) and Salmonella are food-borne pathogens commonly associated with beef, and reliable methods are needed to determine their prevalence in beef and to ensure food safety. Retail ground beef was tested for the presence of E. coli O157:H7, STEC serogroups O26, O45, O103, O111, O121, and O145, and Salmonella using the DuPont™ BAX® system method. Ground beef (325 g) samples were enriched in 1.5 L of TSB with 2 mg/L novobiocin at 42°C for 18 h, and then evaluated using the BAX® System real-time PCR assays for E. coli O157:H7 and STEC suite, and the BAX® System standard PCR assays for E. coli O157:H7 MP and Salmonella. Samples positive for STEC target genes by the BAX® System assays were subjected to immunomagnetic separation (IMS) and plating onto modified Rainbow Agar O157. Enrichments that were PCR positive for Salmonella were inoculated into RV broth, incubated for 18 h at 42°C, and then plated onto XLT-4 agar. Presumptive positive STEC and Salmonella colonies were confirmed using the BAX® System assays. Results of the BAX® System STEC assays showed 20/308 (6.5%) of samples positive for both the Shiga toxin (stx) and intimin (eae) genes; 4 (1.3%) for stx, eae, and O26; 1 (0.3%) for stx, eae, and O45; 3 (1%) for stx, eae, and O103; and 1 (0.3%) for stx, eae, and O145. There were also 3 samples positive for stx, eae, and more than one STEC serogroup. Three (1.0%) of the samples were positive using the BAX® System real-time E. coli O157:H7 assay, and 28 (9.1%) were positive using the BAX® System Salmonella assay. STEC O103 and E. coli O157:H7 were isolated from 2/6 and 2/3 PCR positive samples, respectively. Salmonella isolates were recovered and confirmed from 27 of the 28 Salmonella PCR positive samples, and a portion of the isolates were serotyped and antibiotic resistance profiles determined. Results demonstrate that the BAX® System assays are effective for detecting STEC and Salmonella in beef.

  13. Molecular characterisation of human Shiga toxin-producing Escherichia coli O26 strains: results of an outbreak investigation, Romania, February to August 2016.

    Science.gov (United States)

    Usein, Codruţa-Romaniţa; Ciontea, Adriana Simona; Militaru, Cornelia Mãdãlina; Condei, Maria; Dinu, Sorin; Oprea, Mihaela; Cristea, Daniela; Michelacci, Valeria; Scavia, Gaia; Zota, Lavinia Cipriana; Zaharia, Alina; Morabito, Stefano

    2017-11-01

    IntroductionAt the beginning of 2016, an increase in paediatric haemolytic uremic syndrome (HUS) cases was observed in Romania. The microbiological investigations allowed isolation of Shiga toxin-producing Escherichia coli (STEC) O26 as the causative agent from most cases. Methods: An enhanced national surveillance of HUS and severe diarrhoea was established across the country following the identification of the first cases and was carried out until August 2016. A total of 15 strains were isolated from 10 HUS and five diarrhoea cases. Strains were characterised by virulence markers (i.e. stx type/subtype, eae , ehxA genes), phylogroup, genetic relatedness and clonality using PCR-based assays, PFGE and multilocus sequence typing (MLST). The first six strains were further characterised by whole genome sequencing (WGS). Results: Five PCR-defined genotypes were distinguished. All strains from HUS cases harboured stx2a and eae , with or without stx1a , while strains from diarrhoea cases carried exclusively stx1a and eae genes. PFGE resolved strains into multiple pulsotypes, compatible with a certain geographic segregation of the cases, and strains were assigned to phylogroup B1 and sequence type (ST) 21. WGS confirmed the results of conventional molecular methods, brought evidence of O26:H11 serotype, and complemented the virulence profiles. Discussion/conclusion: This first description of STEC O26 strains from cases in Romania showed that the isolates belonged to a diverse population. The virulence content of most strains highlighted a high risk for severe outcome in infected patients. Improving the national surveillance strategy for STEC infections in Romania needs to be further considered.

  14. Prevalence and behavior of multidrug-resistant shiga toxin-producing Escherichia coli, enteropathogenic E. coli and enterotoxigenic E. coli on coriander.

    Science.gov (United States)

    Gómez-Aldapa, Carlos A; Segovia-Cruz, Jesús A; Cerna-Cortes, Jorge F; Rangel-Vargas, Esmeralda; Salas-Rangel, Laura P; Gutiérrez-Alcántara, Eduardo J; Castro-Rosas, Javier

    2016-10-01

    The prevalence and behavior of multidrug-resistant diarrheagenic Escherichia coli pathotypes on coriander was determined. One hundred coriander samples were collected from markets. Generic E. coli were determined using the most probable number procedure. Diarrheagenic E. coli pathotypes (DEPs) were identified using two multiplex polymerase chain reaction procedures. Susceptibility to sixteen antibiotics was tested for the isolated DEPs strains by standard test. The behavior of multidrug-resistant DEPs isolated from coriander was determined on coriander leaves and chopped coriander at 25°± 2 °C and 3°± 2 °C. Generic E. coli and DEPs were identified, respectively, in 43 and 7% of samples. Nine DEPs strains were isolated from positive coriander samples. The identified DEPs included Shiga toxin-producing E. coli (STEC, 4%) enterotoxigenic E. coli (ETEC, 2%) and enteropathogenic E. coli (EPEC, 1%). All isolated DEPs strains exhibited multi-resistance to antibiotics. On inoculated coriander leaves stored at 25°± 2 °C or 3°± 2 °C, no growth was observed for multidrug-resistant DEPs strains. However, multidrug-resistant DEPs strains grew in chopped coriander: after 24 h at 25° ± 2 °C, DEPs strains had grown to approximately 3 log CFU/g. However, at 3°± 2 °C the bacterial growth was inhibited. To the best of our knowledge, this is the first report of the presence and behavior of multidrug-resistant STEC, ETEC and EPEC on coriander and chopped coriander. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Presence of Staphylococcus aureus and Shiga Toxigenic Escherichia coli O157:H7 in Raw Meat in Ağrı, Turkey

    Directory of Open Access Journals (Sweden)

    Naim Deniz Ayaz

    2016-08-01

    Full Text Available Background: Staphylococcus aureus and Shiga toxin producing Escherichia coli O157:H7 (EHEC are significant foodborne pathogens worldwide. While S. aureus can cause mild superficial skin infections or life-threatening bacteremia and endocarditis, as well as toxininduced cases such as toxic shock syndrome; E. coli O157:H7 can cause symptoms from mild diarrhea to severe hemorrhagic colitis (HC, hemolytic uremic syndrome (HUS, thrombotic thrombocytopenic purpura (TTP. Objectives: The objectives of this study were to find out the prevalence and seasonal distribution of S. aureus in 214 frozen raw meat (turkey, chicken and beef and the prevalence of E. coli O157:H7 in 70 raw beef with the characterization of the E. coli O157:H7 isolate by multiplex polymerase chain reaction (PCR. Materials and Methods: For the detection of S. aureus, a total of 214 frozen raw meat samples including 74 turkey meat, 70 chicken meat and 70 beef cuts (approximately 2 × 3 cm cubic parts; and for the detection of E. coli O157:H7, a total of 70 frozen raw beef samples that all were produced from national companies and consumed in Ağrı, Turkey were analyzed. Results: Out of 214 meat samples, 25.7 % (18/70 of the beef, 11.4 % (8/74 of the chicken meat, and 5.4 % (4/70 of the turkey meat samples were contaminated with S. aureus. Out of 70 frozen raw beef samples, only 1 (1.4% was identified as both Shiga toxin 1 and 2producing E. coli O157:H7 by the detection of stx1, stx2, eaeA, hly, and fliCh7 according to multiplex PCR analysis. Conclusion: Our findings demonstrate that occurrence frequency of S. aureus was higher in frozen raw beef than in raw chicken and turkey meat samples. Although the prevalence of E. coli O157:H7 was low in beef, the presence of virulence genes, especially toxin genesrema in a significant public health concern.

  16. Characterization of novel bacteriophage phiC119 capable of lysing multidrug-resistant Shiga toxin-producing Escherichia coli O157:H7

    Directory of Open Access Journals (Sweden)

    Luis Amarillas

    2016-09-01

    Full Text Available Background Shiga toxin-producing Escherichia coli (STEC is one of the most common and widely distributed foodborne pathogens that has been frequently implicated in gastrointestinal and urinary tract infections. Moreover, high rates of multiple antibiotic-resistant E. coli strains have been reported worldwide. Due to the emergence of antibiotic-resistant strains, bacteriophages are considered an attractive alternative to biocontrol pathogenic bacteria. Characterization is a preliminary step towards designing a phage for biocontrol. Methods In this study, we describe the characterization of a bacteriophage designated phiC119, which can infect and lyse several multidrug-resistant STEC strains and some Salmonella strains. The phage genome was screened to detect the stx-genes using PCR, morphological analysis, host range was determined, and genome sequencing were carried out, as well as an analysis of the cohesive ends and identification of the type of genetic material through enzymatic digestion of the genome. Results Analysis of the bacteriophage particles by transmission electron microscopy showed that it had an icosahedral head and a long tail, characteristic of the family Siphoviridae. The phage exhibits broad host range against multidrug-resistant and highly virulent E. coli isolates. One-step growth experiments revealed that the phiC119 phage presented a large burst size (210 PFU/cell and a latent period of 20 min. Based on genomic analysis, the phage contains a linear double-stranded DNA genome with a size of 47,319 bp. The phage encodes 75 putative proteins, but lysogeny and virulence genes were not found in the phiC119 genome. Conclusion These results suggest that phage phiC119 may be a good biological control agent. However, further studies are required to ensure its control of STEC and to confirm the safety of phage use.

  17. The evolutionary divergence of Shiga toxin-producing Escherichia coli is reflected in clustered regularly interspaced short palindromic repeat (CRISPR) spacer composition.

    Science.gov (United States)

    Yin, Shuang; Jensen, Mark A; Bai, Jiawei; Debroy, Chitrita; Barrangou, Rodolphe; Dudley, Edward G

    2013-09-01

    The Shiga toxin-producing Escherichia coli (STEC) strains, including those of O157:H7 and the "big six" serogroups (i.e., serogroups O26, O45, O103, O111, O121, and O145), are a group of pathogens designated food adulterants in the United States. The relatively conserved nature of clustered regularly interspaced short palindromic repeats (CRISPRs) in phylogenetically related E. coli strains makes them potential subtyping markers for STEC detection, and a quantitative PCR (qPCR)-based assay was previously developed for O26:H11, O45:H2, O103:H2, O111:H8, O121:H19, O145:H28, and O157:H7 isolates. To better evaluate the sensitivity and specificity of this qPCR method, the CRISPR loci of 252 O157 and big-six STEC isolates were sequenced and analyzed along with 563 CRISPR1 and 624 CRISPR2 sequences available in GenBank. General conservation of spacer content and order was observed within each O157 and big-six serogroup, validating the qPCR method. Meanwhile, it was found that spacer deletion, the presence of an insertion sequence, and distinct alleles within a serogroup are sources of false-negative reactions. Conservation of CRISPR arrays among isolates expressing the same flagellar antigen, specifically, H7, H2, and H11, suggested that these isolates share an ancestor and provided an explanation for the false positives previously observed in the qPCR results. An analysis of spacer distribution across E. coli strains provided limited evidence for temporal spacer acquisition. Conversely, comparison of CRISPR sequences between strains along the stepwise evolution of O157:H7 from its O55:H7 ancestor revealed that, over this ∼7,000-year span, spacer deletion was the primary force generating CRISPR diversity.

  18. Feasibility study on production of a matrix reference material for cyanobacterial toxins.

    Science.gov (United States)

    Hollingdale, Christie; Thomas, Krista; Lewis, Nancy; Békri, Khalida; McCarron, Pearse; Quilliam, Michael A

    2015-07-01

    The worldwide increase in cyanobacterial contamination of freshwater lakes and rivers is of great concern as many cyanobacteria produce potent hepatotoxins and neurotoxins (cyanotoxins). Such toxins pose a threat to aquatic ecosystems, livestock, and drinking water supplies. In addition, dietary supplements prepared from cyanobacteria can pose a risk to consumers if they contain toxins. Analytical monitoring for toxins in the environment and in consumer products is essential for the protection of public health. Reference materials (RMs) are an essential tool for the development and validation of analytical methods and are necessary for ongoing quality control of monitoring operations. Since the availability of appropriate RMs for cyanotoxins has been very limited, the present study was undertaken to examine the feasibility of producing a cyanobacterial matrix RM containing various cyanotoxins. The first step was large-scale culturing of various cyanobacterial cultures that produce anatoxins, microcystins, and cylindrospermopsins. After harvesting, the biomass was lyophilized, blended, homogenized, milled, and bottled. The moisture content and physical characteristics were assessed in order to evaluate the effectiveness of the production process. Toxin levels were measured by liquid chromatography with tandem mass spectrometry and ultraviolet detection. The reference material was found to be homogeneous for toxin content. Stability studies showed no significant degradation of target toxins over a period of 310 days at temperatures up to +40 °C except for the anatoxin-a, which showed some degradation at +40 °C. These results show that a fit-for-purpose matrix RM for cyanotoxins can be prepared using the processes and techniques applied in this work.

  19. The Regulation of Expression of the Stx2d Toxins in Shiga Toxin-producing Escherichia coli O91:H21 Strain B2F1

    Science.gov (United States)

    2002-01-01

    done by Edda Twiddy). The mutants were also transduced with bacteriophage 933W to assess cytotoxicity in the DH5α mutants of a related toxin gene in...amounts of toxin antigen produced by DH5α with the levels produced by the mutants (with the assistance of Edda Twiddy). Dot blots were 84

  20. Production of T-2 toxin and deoxynivalenol in the presence of different disinfectants

    Directory of Open Access Journals (Sweden)

    Dana Hrubošová

    2015-03-01

    Full Text Available The aim of the work was to examine the effect of different disinfectants on production trichothecenes (especially of T-2 toxin and deoxynivalenol. Lipophilicity, chemical structure, the presence of bioactive groups and functional groups in their structure modifies biological activity and toxic potency of trichothecenes. For this reason, limits have been established designating maximum levels of mycotoxins in cereals while maintaining proper growing practices. Appropriate nutritive media were prepared with different concentration of tested disinfectants (Desanal A  plus, ProCura spray and Guaa-Pool and were inoculated using  Fusarium strains. The density of  Fusarium was 105 spores per mililitre. Nutrient media was cultivated at 15 °C and 25 °C for seven days. The strains of Fusarium graminearum CCM F-683 and Fusarium species (isolated from barley produced quantities of deoxynivalenol. Fusarium poae CCM F-584 and Fusarium species (isolated from malthouse air produced quantities of T-2 toxin. Desanal A plus prevented Fusarium growth and production of T-2 toxin and deoxynivalenol at the concentration 10%. It is an alkaline disinfectant on the basis of active chlorine and the surfactant that contains ˂5% of NaClO. ProCura spray at the concentration 0.6% proved to be very effective. This disinfectant contains 35% of propan-1-ol and 25% of propan-2-ol.  Guaa-Pool at the concentration 0.004% proved to be very effective. It is a polymeric disinfectant with anion surface-acting agent and it contains ˂0.9% of polyhexamethylene guanidine hydrochloride and ˂0.2% of alkyl (C12-C16 dimethylbenzyl ammonium chloride. Lower contentration of  disinfectants that  not prevented growth of Fusarium caused higher production  of T-2 toxin and deoxynivalenol. The contents of T-2 toxin and deoxynivalenol were analyzed by enzyme-linked immunosorbent assay (ELISA using commercially produced kits (Agra Quant® Deoxynivalenol Test kit and Agra Quant® T-2 toxin

  1. Climate change impacts on natural toxins in food production systems, exemplified by deoxynivalenol in wheat and diarrhetic shellfish toxins

    DEFF Research Database (Denmark)

    van der Fels-Klerx, H J; Olesen, Jørgen E; Naustvoll, L-J

    2012-01-01

    Climate change is expected to affect food and feed safety, including the occurrence of natural toxins in primary crop and seafood production; however, to date, quantitative estimates are scarce. This study aimed to estimate the impact of climate change effects on mycotoxin contamination of cereal...... on food safety hazards, rather than median or average values only. Furthermore, it is recommended to closely monitor levels of mycotoxins and marine biotoxins in the future, in particular related to risky situations associated with favourable climatic conditions for toxin producing organisms...

  2. Rare codons effect on expression of recombinant gene cassette in Escherichia coli BL21(DE3

    Directory of Open Access Journals (Sweden)

    Aghil Esmaeili-Bandboni

    2017-11-01

    Full Text Available Objective: To demonstrate the sensitivity of expression of fusion genes to existence of a large number of rare codons in recombinant gene sequenced. Methods: Primers for amplification of cholera toxin B, Shiga toxin B and gfp genes were designed by Primer3 software and synthesized. All of these 3 genes were cloned. Then the genes were fused together by restriction sites and enzymatic method. Two linkers were used as a flexible bridge in connection of these genes. Results: Cloning and fusion of cholera toxin B, Shiga toxin B and gfp genes were done correctly. After that, expression of the recombinant gene construction was surveyed. Conclusions: According to what was seen, because of the accumulation of 12 rare codons of Shiga toxin B and 19 rare codons of cholera toxin B in this gene cassette, the expression of the recombinant gene cassette, in Escherichia coli BL21, failed.

  3. Genetic characterization of Shiga toxin-producing Escherichia coli (STEC) and atypical enteropathogenic Escherichia coli (EPEC) isolates from goat's milk and goat farm environment.

    Science.gov (United States)

    Álvarez-Suárez, María-Elena; Otero, Andrés; García-López, María-Luisa; Dahbi, Ghizlane; Blanco, Miguel; Mora, Azucena; Blanco, Jorge; Santos, Jesús A

    2016-11-07

    The aim of this study was to characterize a collection of 44 Shiga toxin-producing (STEC) and enteropathogenic Escherichia coli (EPEC) isolated from goat milk and goat farm environment. Of the 19 STEC isolates, five (26.3%) carried the stx1 gene, four (21.1%) the stx2 gene and 10 (52.6%) presented both stx genes. Six (31.6%) STEC strains were eae-positive and belonged to serotypes related to severe human disease (O157:H7 and O5:HNM). Another seven STEC strains were of serotype O146:H21 and three of serotype O166:H28, also linked to human disease. The STEC strains isolated from goat milk were of serotypes potentially pathogenic for humans. All the 25 EPEC isolates were considered atypical (aEPEC) and one aEPEC strain was of serotype O26:H11, a serotype frequently isolated in children with diarrhea. Multilocus sequence typing (MLST) was carried out with seven housekeeping genes and 23 sequence types (ST) were detected, 14 of them newly described. Twelve STs grouped STEC isolates and 11 STs grouped EPEC isolates. Genetic typing by pulsed field gel electrophoresis (PFGE) resulted in 38 patterns which grouped in 10 clusters. Well-defined groups were also observed for strains of pathogenic serotypes. In conclusion, strains of STEC and aEPEC belonging to serotypes related to severe human disease have been detected in goat milk and the goat farm environment. Ruminants are an important reservoir of STEC strains and the role of these animals as carriers of other pathogenic types of E. coli seems to be an emerging concern. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Evidence of non-O157 Shiga toxin-producing Escherichia coli in the feces of meat goats at a U.S. slaughter plant.

    Science.gov (United States)

    Jacob, M E; Foster, D M; Rogers, A T; Balcomb, C C; Shi, X; Nagaraja, T G

    2013-09-01

    Shiga toxin-producing Escherichia coli (STEC) are important human pathogens, and attention to non-O157 serogroups has increased in recent years. Although cattle are normally considered the primary reservoir for STEC, recent illnesses associated with goat contact have indicated that these animals are important potential reservoirs for the organisms. The prevalence of STEC, particularly non-O157 serogroups, in U.S. goats has not been well described. Our objective was to determine the prevalence of six major non-O157 STEC serogroups in the feces of meat goats. Rectal contents from 296 goats were collected postevisceration at a slaughter plant in the southeastern United States over 9 days during a 12-week period from August through October 2012. Samples were enriched in E. coli broth, and DNA was extracted and used as template in an 11-gene multiplex PCR that detected six non-O157 serogroups (O26, O45, O103, O121, O111, and O145) and virulence genes. Samples were considered positive when at least one non-O157 STEC serotype was present with either stx₁ or stx₂. All six non-O157 serogroups were detected by PCR in our samples, and 14.5% of samples were positive for at least one serogroup. Prevalence of O26 was highest, with 6.4% of goat fecal samples positive. The prevalence of O45 was 3.4%, O103 was 4.4%, O111 was 4.1%, O121 was 1.4%, and O145 was 3.0%. Twenty-two (7.4%) of 296 fecal samples had more than one non-O157 serogroup detected in the feces. Two samples had evidence of three non-O157 STEC serogroups. Goats appear to be an important reservoir for non-O157 STEC, and further work to understand the characteristics, epidemiology, and ecology of STEC in these animals is warranted.

  5. Canonical Single Nucleotide Polymorphisms (SNPs for High-Resolution Subtyping of Shiga-Toxin Producing Escherichia coli (STEC O157:H7.

    Directory of Open Access Journals (Sweden)

    Sean M Griffing

    Full Text Available The objective of this study was to develop a canonical, parsimoniously-informative SNP panel for subtyping Shiga-toxin producing Escherichia coli (STEC O157:H7 that would be consistent with epidemiological, PFGE, and MLVA clustering of human specimens. Our group had previously identified 906 putative discriminatory SNPs, which were pared down to 391 SNPs based on their prevalence in a test set. The 391 SNPs were screened using a high-throughput form of TaqMan PCR against a set of clinical isolates that represent the most diverse collection of O157:H7 isolates from outbreaks and sporadic cases examined to date. Another 30 SNPs identified by others were also screened using the same method. Two additional targets were tested using standard TaqMan PCR endpoint analysis. These 423 SNPs were reduced to a 32 SNP panel with the almost the same discriminatory value. While the panel partitioned our diverse set of isolates in a manner that was consistent with epidemiological data and PFGE and MLVA phylogenies, it resulted in fewer subtypes than either existing method and insufficient epidemiological resolution in 10 of 47 clusters. Therefore, another round of SNP discovery was undertaken using comparative genomic resequencing of pooled DNA from the 10 clusters with insufficient resolution. This process identified 4,040 potential SNPs and suggested one of the ten clusters was incorrectly grouped. After its removal, there were 2,878 SNPs, of which only 63 were previously identified and 438 occurred across multiple clusters. Among highly clonal bacteria like STEC O157:H7, linkage disequilibrium greatly limits the number of parsimoniously informative SNPs. Therefore, it is perhaps unsurprising that our panel accounted for the potential discriminatory value of numerous other SNPs reported in the literature. We concluded published O157:H7 SNPs are insufficient for effective epidemiological subtyping. However, the 438 multi-cluster SNPs we identified may provide

  6. Presence of non-O157 Shiga toxin-producing Escherichia coli, enterotoxigenic E. coli, enteropathogenic E. coli and Salmonella in fresh beetroot (Beta vulgaris L.) juice from public markets in Mexico.

    Science.gov (United States)

    Gómez-Aldapa, Carlos A; Rangel-Vargas, Esmeralda; Bautista-De León, Haydee; Castro-Rosas, Javier

    2014-10-01

    Unpasteurized juice has been associated with foodborne illness outbreaks for many years. Beetroot is a vegetable grown all over the world in temperate areas. In Mexico beetroot is consumed cooked in salads or raw as fresh unpasteurized juices. No data about the microbiological quality or safety of unpasteurized beetroot juices are available. Indicator bacteria, diarrheagenic Escherichia coli pathotypes (DEP) and Salmonella frequencies were determined for fresh unpasteurized beetroot juice from restaurants. One hundred unpasteurized beetroot juice samples were collected from public markets in Pachuca, Mexico. Frequencies in these samples were 100%, 75%, 53%, 9% and 4% of positive samples, for coliform bacteria, fecal coliforms, E. coli, DEP and Salmonella, respectively. Identified DEP included enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC) and non-O157 Shiga toxin-producing E. coli (STEC). Identified Salmonella serotypes included Typhimurium and Enteritidis. This is the first report of microbiological quality and atypical EPEC, ETEC, non-O157 STEC and Salmonella isolation from fresh raw beetroot juice in Mexico. Fresh raw beetroot juice from markets is very probably an important factor contributing to the endemicity of atypical EPEC, ETEC, non-O157 STEC and Salmonella-related gastroenteritis in Mexico. © 2014 Society of Chemical Industry.

  7. Effect of gamma irradiation on genetic changes and myco toxin production by fungi isolated from corn grains

    International Nuclear Information System (INIS)

    Abou-Sereih, N.A.; Sahab, A.F.; El-Fiki, A.A.M.

    1998-01-01

    Three important fungi and the major myco toxins were isolated from different food and feed products. The effect of gamma irradiation on genome structures and its relation to the production of myco toxin were also investigated. The percentage of auxotrophic mutants and its relation to the production of myco toxin, when the strains were treated with gamma irradiation (o.1, 0.2, 0.3 and 0.4 kgy), were determined. In general, gamma radiation resulted in high percentage of mutation at 0.4 kgy and the production of mycotoxin decreased greatly or inhibited by 0.1, 0.2, 0.3 and 0.4 kgy due to change in the genetic construction of genes responsible for mycotoxin production

  8. In vitro evaluation of the effect of linezolid and levofloxacin on Bacillus anthracis toxin production, spore formation and cell growth.

    Science.gov (United States)

    Head, Breanne M; Alfa, Michelle; Sitar, Daniel S; Rubinstein, Ethan; Meyers, Adrienne F A

    2017-02-01

    Owing to its ability to form spores and toxins, Bacillus anthracis is considered a bioterror agent. Although current therapeutic strategies can be effective, treatment does not prevent sporulation and toxin production. To quantify the combined effect of a protein synthesis inhibitor and a bactericidal agent on B. anthracis toxin production, sporulation and cell growth. Susceptibility and synergy titrations were conducted on B. anthracis Sterne and 03-0191 strains using linezolid and levofloxacin. The effect of antibiotic exposure on cell viability was evaluated using a continuous medium replacement model. In vitro static models were used to study the effect of linezolid and levofloxacin on sporulation and toxin production. Spores were quantified using the heat shock method. Toxin was quantified via commercial ELISA. Synergy titrations indicated that the combination was synergistic or indifferent; however, in all models antagonism was observed. In the spore model, linezolid resulted in the lowest sporulation rates, while combination therapy resulted in the highest. In the toxin model, linezolid prevented toxin production altogether. This study advances our understanding of the effects of combination therapy on B. anthracis infection. Used alone, linezolid therapy abolishes toxin production and reduces sporulation. These results suggest that studies using a step-wise approach using linezolid initially to stop sporulation and toxin production followed by levofloxacin to rapidly kill vegetative B. anthracis can be recommended. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  9. Effect of rifampicin and gentamicin on Shiga toxin 2 expression level and the SOS response in Escherichia coli O104:H4.

    Science.gov (United States)

    Fadlallah, Sukayna M; Rahal, Elias A; Sabra, Ahmad; Kissoyan, Kohar A B; Matar, Ghassan M

    2015-01-01

    A novel pathotype, Shiga toxin-producing Escherichia coli O104:H4, was the cause of a severe outbreak that affected European countries, mainly Germany, in 2011. The effect of different regimens of rifampicin and gentamicin were evaluated to determine possible treatment modes for the novel strain, and to evaluate the SOS response and its effect on toxin release. Pulsed-field gel electrophoresis (PFGE) was performed on the novel E. coli O104:H4 pathotype and two pre-outbreak E. coli O104:H4 CDC strains. Transcript levels of the stx2 and recA gene (SOS response inducer) were evaluated using quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR) in the novel E. coli O104:H4 samples subjected to different regimens of rifampicin and gentamicin. Consequently, reverse passive latex agglutination (RPLA) was used to determine the Stx2 titers in these samples. Western blot was performed to determine the LexA levels (SOS response repressor) in E. coli O104:H4. The efficacy of treatment with antimicrobial agents was assessed in BALB/c mice. The outbreak and pre-outbreak strains are closely related as shown by PFGE, which demonstrated slight genomic differences between the three strains. The transcription level of the stx2 gene in the new pathotype was 1.41- and 1.75-fold that of the 2009 EL-2050 and 2009 EL-2071 pre-outbreak strains, respectively. Moreover, the transcription level of the stx2 gene in the new pathotype was substantially decreased as a result of treatment with the different concentrations of the antimicrobial agents, but was enhanced when the antibiotics were administered at two subinhibitory levels. RPLA data were in accordance with the qRT-PCR results. E. coli O104:H4 exposed to gentamicin at both sub-minimum inhibitory concentration (MIC) levels led to high transcription levels of the recA gene and lack of expression of the LexA protein, implying that the SOS response was activated. Rifampicin at both sub-MIC levels resulted in low

  10. Endoplasmic Reticulum-Targeted Subunit Toxins Provide a New Approach to Rescue Misfolded Mutant Proteins and Revert Cell Models of Genetic Diseases.

    Science.gov (United States)

    Adnan, Humaira; Zhang, Zhenbo; Park, Hyun-Joo; Tailor, Chetankumar; Che, Clare; Kamani, Mustafa; Spitalny, George; Binnington, Beth; Lingwood, Clifford

    2016-01-01

    Many germ line diseases stem from a relatively minor disturbance in mutant protein endoplasmic reticulum (ER) 3D assembly. Chaperones are recruited which, on failure to correct folding, sort the mutant for retrotranslocation and cytosolic proteasomal degradation (ER-associated degradation-ERAD), to initiate/exacerbate deficiency-disease symptoms. Several bacterial (and plant) subunit toxins, retrograde transport to the ER after initial cell surface receptor binding/internalization. The A subunit has evolved to mimic a misfolded protein and hijack the ERAD membrane translocon (dislocon), to effect cytosolic access and cytopathology. We show such toxins compete for ERAD to rescue endogenous misfolded proteins. Cholera toxin or verotoxin (Shiga toxin) containing genetically inactivated (± an N-terminal polyleucine tail) A subunit can, within 2-4 hrs, temporarily increase F508delCFTR protein, the major cystic fibrosis (CF) mutant (5-10x), F508delCFTR Golgi maturation (glucocerobrosidase (GCC) in N370SGCC Gaucher Disease fibroblasts (3x), another ERAD-exacerbated misfiling disease. We identify a new, potentially benign approach to the treatment of certain genetic protein misfolding diseases.

  11. Toxin production by Clostridium Botulinum type B (proteolitic) in radurized raw fish

    International Nuclear Information System (INIS)

    Suhadi, F.

    1978-01-01

    The earliest toxin production by three proteolytic strains of Cl. botulinum type B was determined in irradiated and unirradiated raw fish (Rastrelliger sp., Euthynnus sp., and Scomberomorus sp.) under the storage temperatures of 20, 10, and 5degC. The estimation of maximum storage life was evaluated by an untrained panel on uninoculated fish samples and in parellel the total bacterial counts were also determined. Percentage data of the toxic samples were analyzed according to a fully randomized design involving factorial treatments. In unirradiated samples with inoculum levels of 10 2 -10 6 spores per gram and stored at 20degC, the earliest toxin production was detected after the samples were spoiled. While in irradiated samples toxin were detected before the end of the storage life or after the samples were spoiled, depending on the levels of inoculum. In general, both in unirradiated and irradiated samples inoculated with 10 2 -10 6 spores per gram and stored at 10degC, the earlieast toxin production was detected after the samples were spoiled. While the samples were stored at 5degC, no toxic samples were found up to 30 days of storage when the experiment were terminated. The percentage of toxic samples was shown highly effected by type B strains, fish species, inoculum levels and storage time, when the storage temperature is 20degC. But no significant difference was found after treatment with irradiation doses. In general the interaction effects between those treatments on the percentage of toxic samples showed no significant difference. (author)

  12. Detection of Staphylococcus aureus delta-toxin production by whole-cell MALDI-TOF mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Julie Gagnaire

    Full Text Available The aim of the present study was to detect the Staphylococcus aureus delta-toxin using Whole-Cell (WC Matrix Assisted Laser Desorption Ionization-Time-of-Flight (MALDI-TOF mass spectrometry (MS, correlate delta-toxin expression with accessory gene regulator (agr status, and assess the prevalence of agr deficiency in clinical isolates with and without resistance to methicillin and glycopeptides. The position of the delta-toxin peak in the mass spectrum was identified using purified delta-toxin and isogenic wild type and mutant strains for agr-rnaIII, which encodes delta-toxin. Correlation between delta-toxin production and agr RNAIII expression was assessed by northern blotting. A series of 168 consecutive clinical isolates and 23 unrelated glycopeptide-intermediate S. aureus strains (GISA/heterogeneous GISA were then tested by WC-MALDI-TOF MS. The delta-toxin peak was detected at 3005±5 Thomson, as expected for the naturally formylated delta toxin, or at 3035±5 Thomson for its G10S variant. Multivariate analysis showed that chronicity of S. aureus infection and glycopeptide resistance were significantly associated with delta-toxin deficiency (p = 0.048; CI 95%: 1.01-10.24; p = 0.023; CI 95%: 1.20-12.76, respectively. In conclusion, the S. aureus delta-toxin was identified in the WC-MALDI-TOF MS spectrum generated during routine identification procedures. Consequently, agr status can potentially predict infectious complications and rationalise application of novel virulence factor-based therapies.

  13. Profile of Shiga toxin-producing Escherichia coli strains isolated from dogs and cats and genetic relationships with isolates from cattle, meat and humans.

    Science.gov (United States)

    Bentancor, A; Rumi, M V; Carbonari, C; Gerhardt, E; Larzábal, M; Vilte, D A; Pistone-Creydt, V; Chinen, I; Ibarra, C; Cataldi, A; Mercado, E C

    2012-05-04

    Pets can be reservoirs of Shiga toxin-producing Escherichia coli (STEC) strains. The aim of this study was to examine nine strains belonging to several serotypes (O91:H21, O91:H16, O178:H19, O8:H19, O22:H8, O22:HNT, ONT:H8), previously recovered from cats or dogs. To this end, we assessed a set of additional virulence genes (stx(2) subtype, subAB, ehxA, eae and saa), cytotoxic activity, and genetic relationships with strains isolated from cattle, meat and humans using pulsed-field gel electrophoresis (PFGE). Most of the isolates carried the stx(2) and/or stx(2vh-b) sequences, while only the O91:H21 isolate presented the mucus-activatable stx(2d) variant, as confirmed by sequencing the genes of subunits A and B. All the strains showed cytotoxic activity in cultured cells. One of the two O178:H19, selected for its high level of cytotoxicity in Vero cells, showed the ability to cause functional alterations in the human colon mucosa in vitro. None of the strains possessed the subAB, eae or saa genes and only the strains belonging to serotype O8:H19 carried the ehxA gene. The isolates shared 90-100% similarity by PFGE to epidemiologically unrelated strains of the corresponding serotypes recovered from cattle, meat or humans. Our results demonstrate that dogs and cats may have a role in the infection of humans by STEC, probably serving as a vehicle for bovine strains in the cycle of human infection, and thus emphasize the health risks for owners and their families. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Origins of the E. coli strain causing an outbreak of hemolytic-uremic syndrome in Germany

    DEFF Research Database (Denmark)

    Rasko, David A; Webster, Dale R; Sahl, Jason W

    2011-01-01

    A large outbreak of diarrhea and the hemolytic-uremic syndrome caused by an unusual serotype of Shiga-toxin-producing Escherichia coli (O104:H4) began in Germany in May 2011. As of July 22, a large number of cases of diarrhea caused by Shiga-toxin-producing E. coli have been reported--3167 without...... the hemolytic-uremic syndrome (16 deaths) and 908 with the hemolytic-uremic syndrome (34 deaths)--indicating that this strain is notably more virulent than most of the Shiga-toxin-producing E. coli strains. Preliminary genetic characterization of the outbreak strain suggested that, unlike most of these strains......, it should be classified within the enteroaggregative pathotype of E. coli....

  15. Study on potential Clostridium botulinum growth and toxin production in Parma ham

    Directory of Open Access Journals (Sweden)

    Giuseppe Merialdi

    2016-04-01

    Full Text Available The objective of this study was to investigate Clostridium botulinum growth and toxin production in the industrially manufactured Italian Parma ham. The study focuses on the Parma ham production phase identified as maximum risk to C. botulinum proliferation, i.e. the transition from cold phase (salting and resting to a phase carried out at temperature between 15 and 23°C (drying. A preliminary in vitro test was carried out in order to verify the capability of 6 C. botulinum strains (1 type A, 4 type B, and 1 type E strains to grow in conditions of temperature, pH and NaCl concentration comparable to those of the beginning stage of ham drying. Five C. botulinum strains grew at 20°C and pH 6, four strains produced toxin when inoculated at a concentration equal to 103 cfu/mL at NaCl concentration of 4%, while when the inoculum concentration was 10 cfu/mL, NaCl concentration of 3% resulted the toxin-genesis limiting factor. An experimental contamination with a mixture of the 5 C. botulinum strains selected by the preliminary in vitro test was performed on 9 thighs inoculated at the end of the resting phase. The study was designed to evaluate the potential growth and toxin production in extremely favourable conditions for the bacterium. Type B proteolytic C. botulinum toxin was produced after 14 days of incubation at 20°C in 2 thighs characterised by high weight, low number of days of resting and anomalous physiochemical characteristics [one for very low NaCl concentration (1.59%, the other for elevated pH (6.27 and both for high water activity values (>0.970]. The results of this research confirm that the cold resting step is a critical phase in the production process of Parma ham for the investigated hazard. Based on the present study, the long resting phase adopted in the manufacturing of Parma ham is proven effective to prevent the growth of C. botulinum, an event which could not otherwise be excluded if the hams were processed under less

  16. Implementation of Whole Genome Sequencing (WGS for Identification and Characterization of Shiga Toxin-Producing Escherichia coli (STEC in the United States

    Directory of Open Access Journals (Sweden)

    Rebecca L Lindsey

    2016-05-01

    Full Text Available Shiga toxin-producing Escherichia coli (STEC is an important foodborne pathogen capable of causing severe disease in humans. Rapid and accurate identification and characterization techniques are essential during outbreak investigations. Current methods for characterization of STEC are expensive and time-consuming. With the advent of rapid and cheap whole genome sequencing (WGS benchtop sequencers, the potential exists to replace traditional workflows with WGS. The aim of this study was to validate tools to do reference identification and characterization from WGS for STEC in a single workflow within an easy to use commercially available software platform. Publically available serotype, virulence, and antimicrobial resistance databases were downloaded from the Center for Genomic Epidemiology (CGE (www.genomicepidemiology.org and integrated into a genotyping plug-in with in silico PCR tools to confirm some of the virulence genes detected from WGS data. Additionally, down sampling experiments on the WGS sequence data were performed to determine a threshold for sequence coverage needed to accurately predict serotype and virulence genes using the established workflow. The serotype database was tested on a total of 228 genomes and correctly predicted from WGS for 96.1% of O serogroups and 96.5% of H serogroups identified by conventional testing techniques. A total of 59 genomes were evaluated to determine the threshold of coverage to detect the different WGS targets, 40 were evaluated for serotype and virulence gene detection and 19 for the stx gene subtypes. For serotype, 95% of the O and 100% of the H serogroups were detected at > 40x and ≥ 30x coverage, respectively. For virulence targets and stx gene subtypes, nearly all genes were detected at > 40x, though some targets were 100% detectable from genomes with coverage ≥20x. The resistance detection tool was 97% concordant with phenotypic testing results. With isolates sequenced to > 40x

  17. Implementation of Whole Genome Sequencing (WGS) for Identification and Characterization of Shiga Toxin-Producing Escherichia coli (STEC) in the United States

    Science.gov (United States)

    Lindsey, Rebecca L.; Pouseele, Hannes; Chen, Jessica C.; Strockbine, Nancy A.; Carleton, Heather A.

    2016-01-01

    Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen capable of causing severe disease in humans. Rapid and accurate identification and characterization techniques are essential during outbreak investigations. Current methods for characterization of STEC are expensive and time-consuming. With the advent of rapid and cheap whole genome sequencing (WGS) benchtop sequencers, the potential exists to replace traditional workflows with WGS. The aim of this study was to validate tools to do reference identification and characterization from WGS for STEC in a single workflow within an easy to use commercially available software platform. Publically available serotype, virulence, and antimicrobial resistance databases were downloaded from the Center for Genomic Epidemiology (CGE) (www.genomicepidemiology.org) and integrated into a genotyping plug-in with in silico PCR tools to confirm some of the virulence genes detected from WGS data. Additionally, down sampling experiments on the WGS sequence data were performed to determine a threshold for sequence coverage needed to accurately predict serotype and virulence genes using the established workflow. The serotype database was tested on a total of 228 genomes and correctly predicted from WGS for 96.1% of O serogroups and 96.5% of H serogroups identified by conventional testing techniques. A total of 59 genomes were evaluated to determine the threshold of coverage to detect the different WGS targets, 40 were evaluated for serotype and virulence gene detection and 19 for the stx gene subtypes. For serotype, 95% of the O and 100% of the H serogroups were detected at > 40x and ≥ 30x coverage, respectively. For virulence targets and stx gene subtypes, nearly all genes were detected at > 40x, though some targets were 100% detectable from genomes with coverage ≥20x. The resistance detection tool was 97% concordant with phenotypic testing results. With isolates sequenced to > 40x coverage, the different

  18. Phage Types and Genotypes of Shiga Toxin-Producing Escherichia coli O157:H7 Isolates from Humans and Animals in Spain: Identification and Characterization of Two Predominating Phage Types (PT2 and PT8)

    Science.gov (United States)

    Mora, Azucena; Blanco, Miguel; Blanco, Jesús E.; Alonso, M. Pilar; Dhabi, Ghizlane; Thomson-Carter, Fiona; Usera, Miguel A.; Bartolomé, Rosa; Prats, Guillermo; Blanco, Jorge

    2004-01-01

    Phage typing and DNA macrorestriction fragment analysis by pulsed-field electrophoresis (PFGE) were used for the epidemiological subtyping of a collection of Shiga toxin-producing Escherichia coli (STEC) O157:H7 strains isolated in Spain between 1980 and 1999. Phage typing distinguished a total of 18 phage types among 171 strains isolated from different sources (67 humans, 82 bovines, 12 ovines, and 10 beef products). However, five phage types, phage type 2 (PT2; 42 strains), PT8 (33 strains), PT14 (14 strains), PT21/28 (11 strains), and PT54 (16 strains), accounted for 68% of the study isolates. PT2 and PT8 were the most frequently found among strains from both humans (51%) and bovines (46%). Interestingly, we detected a significant association between PT2 and PT14 and the presence of acute pathologies. A group of 108 of the 171 strains were analyzed by PFGE, and 53 distinct XbaI macrorestriction patterns were identified, with 38 strains exhibiting unique PFGE patterns. In contrast, phage typing identified 15 different phage types. A total of 66 phage type-PFGE subtype combinations were identified among the 108 strains. PFGE subtyping differentiated between unrelated strains that exhibited the same phage type. The most common phage type-PFGE pattern combinations were PT2-PFGE type 1 (1 human and 11 bovine strains), PT8-PFGE type 8 (2 human, 6 bovine, and 1 beef product strains), PT2-PFGE subtype 4A (1 human, 3 bovine, and 1 beef product strains). Nine (29%) of 31 human strains showed phage type-PFGE pattern combinations that were detected among the bovine strains included in this study, and 26 (38%) of 68 bovine strains produced phage type-PFGE pattern combinations observed among human strains included in this study, confirming that cattle are a major reservoir of strains pathogenic for humans. PT2 and PT8 strains formed two groups which differed from each other in their motilities, stx genotypes, PFGE patterns, and the severity of the illnesses that they caused

  19. Validación de una técnica de PCR múltiple para la detección de Escherichia coli productor de toxina Shiga Validation of a multiplex PCR for detection of Shiga toxin-producing Escherichia coli

    Directory of Open Access Journals (Sweden)

    G.A. Leotta

    2005-03-01

    Full Text Available La infección por Escherichia coli productor de toxina Shiga (STEC es causa de diarrea con o sin sangre, colitis hemorrágica y síndrome urémico hemolítico (SUH en humanos. El objetivo de este trabajo fue validar una técnica de PCR múltiple para el diagnóstico de STEC basado en la detección de los genes stx1, stx2 y rfbO157. La validación de la técnica se realizó en dos laboratorios independientes, en forma paralela. Se determinó rango de trabajo, selectividad y robustez. Se evaluó el desempeño de la técnica al combinar distintas concentraciones de dos cepas con diferentes factores de virulencia. El rango de trabajo dependió de la cepa analizada, los valores máximos y mínimos fueron 6,6 x 107 y 1,0 x 104 UFC/50 µl. El límite de detección fue de 1,0 x 104 UFC/50 µl y el límite de corte de 1,0 x 105 UFC/50 µl. La robustez fue óptima al modificar diferentes variables. Se obtuvo 100% de inclusividad, exclusividad, precisión analítica, valor predictivo positivo y valor predictivo negativo. No se observó interferencia al combinar distintas concentraciones de los factores de virulencia blanco de la reacción. La técnica validada es una alternativa apropiada para la detección y confirmación de STEC O157 y no-O157 a partir de cultivos bacterianos.Shiga toxin-producing Escherichia coli (STEC cause non-bloody or bloody diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS in humans. The aim of the present study was to validate a multiplex PCR for the STEC diagnosis based on the detection of stx1, stx2 and rfbO157 genes. The multiplex PCR validation was carried out in two independent laboratories in a parallel way. Work range, selectivity and robustness were established. The PCR performance was evaluated using different concentrations of two STEC strains harboring different target genes. The work range depended on the strain analyzed, the maximum and the minimum values were 6.6 x 107 and 1.0 x 104 CFU/50 µl. The

  20. Climate change impacts on natural toxins in food production systems, exemplified by deoxynivalenol in wheat and diarrhetic shellfish toxins

    NARCIS (Netherlands)

    Fels-Klerx, van der H.J.; Olesen, J.E.; Naustvoll, L.J.; Friocourt, Y.; Mengelers, M.J.B.; Christensen, J.H.

    2012-01-01

    Climate change is expected to affect food and feed safety, including the occurrence of natural toxins in primary crop and seafood production; however, to date, quantitative estimates are scarce. This study aimed to estimate the impact of climate change effects on mycotoxin contamination of cereal

  1. Report on the survey for working out the new energy vision of the Shiga village area; 2001 nendo Shiga mura chiiki shin energy vision sakutei chosa hokokusho

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2002-03-01

    For the purpose of promoting the introduction of new energy and enhancing the awareness of it at Shiga Village, Nagano Prefecture, an investigational study was conducted of the energy demand in the village, existence amount of new energy, project for new energy introduction, etc., and a vision was worked out. The energy consumption amount in FY 2000 in Shiga Village was approximately 50.2 billion kcal and was broken down into 54.7% of petroleum, 38.3% of electric power, 6.3% of LPG and 0.75% of others. As the project for new energy introduction, the following were studied: installation of photovoltaic power generation panels to the new office under construction, introduction of photovoltaic power/wind power hybrid power generation to Klein Garten Club House, installation of the solar system to hot spring facilities in the village, project on low-temperature wood drying by smoking using sawdust/wood chips, methanol production using thinnings as raw materials and the use for automobile fuel/cogeneration, methane fermentation from animal feces/house garbage and the use for cogeneration, etc. (NEDO)

  2. Prevalence of foodborne pathogens in grilled chicken from street vendors and retail outlets in Reynosa, Tamaulipas, Mexico.

    Science.gov (United States)

    Díaz-López, A; Cantú-Ramírez, R C; Garza-González, E; Ruiz-Tolentino, L; Tellez-Luis, S J; Rivera, G; Bocanegra-García, V

    2011-08-01

    We analyzed a total of 70 grilled chicken samples bought randomly from street vendors and retail outlets in the city of Reynosa, Mexico, to determine the prevalence of Escherichia coli (Shiga toxin producing and enterotoxin producing), Salmonella spp., Staphylococcus aureus, Listeria spp., and Campylobacter spp. using microbiological methods and PCR detection of bacterial sequences. Of the 70 samples, 27 (38.5%) were from retail outlets and 43 (61.4%) from street vendors. All specimens were negative by both microbiological and molecular methods for Listeria monocytogenes, Shiga toxin 2 of Shiga toxin-producing E. coli, lt of enterotoxin-producing E. coli, and st enterotoxin, and all were negative for Salmonella spp. and Campylobacter jejuni by PCR. Of the samples studied, 49 (70%) had undetectable levels of the foodborne pathogens studied with the methods used. In the remaining 21 (30%) specimens, at least one pathogen was isolated or detected, with E. coli being the pathogen most frequently isolated and with two samples bearing the hlyA gene. We found no statistical difference in bacterial prevalence between retail and street vendor samples. The presence of pathogens in grilled chicken is an important public health risk because of the great demand for and daily consumption of this product in this region.

  3. Molecular characterization of Escherichia coli recovered from traditional milk products in Kashan, Iran

    Directory of Open Access Journals (Sweden)

    Farhad Sharafati Chaleshtori

    2017-10-01

    Full Text Available Aim: Shiga toxigenic Escherichia coli (STEC strains as emerging groups of foodborne pathogens are responsible for most foodborne illnesses. The aim of this study was to determine the antibiotic resistance pattern in STEC isolated from traditional milk products and their molecular characterization. Materials and Methods: A total of 116 samples were randomly purchased from local markets in Kashan, Iran, and evaluated for the occurrence of STEC by culturing and molecular methods. The antibiotic resistance of obtained isolates was determined by Kirby Bauer method. Furthermore, isolates were assayed for the presence of Shiga toxins (stx1 and stx2 and intimin gene (eae. Results: The incidence of E. coli in 60 ice cream, 30 yoghurt, and 26 cheese samples was 8.33%, 10%, and 11.54%, respectively. The findings showed that 11 out of 11 (100% E. coli had both stx1 and stx2 while eae gene was not found in E. coli isolated of traditional milk products. For E. coli strains carrying stx1 and stx2, highest antibiotic sensitive levels were related to trimethoprim/sulfamethoxazole, norfloxacin, chloramphenicol, and ciprofloxacin, respectively. Conclusion: The results showed relationship between the presence of virulence factors and antimicrobial resistance. These results can be used for further studies on STEC as an emerging foodborne pathogen.

  4. Methods for detecting pathogens in the beef food chain: an overview

    Science.gov (United States)

    The main food-borne pathogens of concern in the beef chain are Shiga toxin-producing Escherichia coli (STEC) and Salmonella. Other pathogens, including Listeria monocytogenes and Campylobacter spp. may also be present and pose contamination concerns in both the cattle production environment and bee...

  5. Occurrence and characterisation of pathogenic Escherichia coli isolated from beef meat imported from South Africa and marketed in united Arab Emirates

    Science.gov (United States)

    Dulaimi, Samar Abdulkareem Khalifa Al; Ibrahim, Nazlina

    2018-04-01

    This study is aimed at determining the presence of Escherichia coli and characterizing the availability of Shiga toxin gene in beef meat samples imported from South Africa. Meat samples (n=47) were randomly collected from meat sections at different supermarkets in the United Arab Emirates (UAE) from the beginning of August 2016 till the end of March 2017. Samples were diluted and inoculated on MacConkey agar and Eosin methylene blue agar (EMB), and the colony forming units (CFU) were recorded. API 20E identification kit was used for biochemical identification of E. coli. Detection of the 16S rRNA and shiga toxin genes by PCR amplification was done. Our results revealed that 14 (29.7%) out of the total 47 samples were positive for E. coli. The bacterial burden of E. coli as determined by colony growth showed variable level of contamination. From the PCR amplification, the shiga toxin gene carried by the E. coli is the Stx2 gene. This study revealed moderately high contamination levels of E. coli in beef samples imported from South Africa and marketed in UAE which mostly carries the shiga toxin gene Stx2.

  6. Draft genome sequences of Escherichia coli O113:H21 strains recovered from a major produce-production region in California

    Science.gov (United States)

    Shiga toxin-producing Escherichia coli is a foodborne and waterborne pathogen and is responsible for outbreaks of human gastroenteritis. This report documents the draft genome sequences of seven O113:H21 strains recovered from livestock, wildlife, and soil samples collected in a major agricultural r...

  7. OI-57, a Genomic Island of Escherichia coli O157, Is Present in Other Seropathotypes of Shiga Toxin-Producing E. coli Associated with Severe Human Disease▿

    Science.gov (United States)

    Imamovic, Lejla; Tozzoli, Rosangela; Michelacci, Valeria; Minelli, Fabio; Marziano, Maria Luisa; Caprioli, Alfredo; Morabito, Stefano

    2010-01-01

    Strains of Shiga toxin-producing Escherichia coli (STEC) are a heterogeneous E. coli group that may cause severe disease in humans. STEC have been categorized into seropathotypes (SPTs) based on their phenotypic and molecular characteristics and the clinical features of the associated diseases. SPTs range from A to E, according to a decreasing rank of pathogenicity. To define the virulence gene asset (“virulome”) characterizing the highly pathogenic SPTs, we used microarray hybridization to compare the whole genomes of STEC belonging to SPTs B, C, and D with that of STEC O157 (SPT A). The presence of the open reading frames (ORFs) associated with SPTs A and B was subsequently investigated by PCR in a larger panel of STEC and in other E. coli strains. A genomic island termed OI-57 was present in SPTs A and B but not in the other SPTs. OI-57 harbors the putative virulence gene adfO, encoding a factor enhancing the adhesivity of STEC O157, and ckf, encoding a putative killing factor for the bacterial cell. PCR analyses showed that OI-57 was present in its entirety in the majority of the STEC genomes examined, indicating that it represents a stable acquisition of the positive clonal lineages. OI-57 was also present in a high proportion of the human enteropathogenic E. coli genomes assayed, suggesting that it could be involved in the attaching-and-effacing colonization of the intestinal mucosa. In conclusion, OI-57 appears to be part of the virulome of pathogenic STEC and further studies are needed to elucidate its role in the pathogenesis of STEC infections. PMID:20823207

  8. Effect of γ-irradiation on F-2 and T-2 toxin production in corn a rice

    International Nuclear Information System (INIS)

    Halasz, A.; Badaway, A.; Sawinsky, J.; Kozma-Kovacs, E.; Beczner, J.

    1989-01-01

    Fusarium graminearum and F. tricinctum were grown on moistened corn and rice. After inoculation the substrates were exposed to γ radiation and the growth rate and mycotoxin production were measured. A delay in mycelium growth and an increase in F-2 and T-2 toxin production occurred after irradiation with 1 and 3 kGy. The maximum F-2 production was 10.7 mg/kg for rice at 3 kGy whereas for T-2 it was 735 μg/kg for rice at 3 kGy. At 9 kGy neither growth nor toxin production could be detected in any inoculated corn and rice substrate. (author). 3 tabs., 12 refs

  9. Efficient mosquitocidal toxin production by Bacillus sphaericus using cheese whey permeate under both submerged and solid state fermentations.

    Science.gov (United States)

    El-Bendary, Magda A; Moharam, Maysa E; Foda, M S

    2008-05-01

    Whey permeate (WP) was used efficiently for production of mosquitocidal toxin by Bacillus sphaericus 2362 (B. sphaericus 2362) and the Egyptian isolate, B. sphaericus 14N1 (B. sphaericus 14N1) under both submerged and solid state fermentation conditions. Under submerged fermentation, high mosquitocidal activity was produced by B. sphaericus 2362 and B. sphaericus 14N1 at 50-100% and 25-70% WP, respectively. Initial pH of WP was a critical factor for toxin production by both tested organisms. The highest toxicity was obtained at initial pH 7. Egyptian isolate, B. sphaericus 14N1 was tested for growth and toxin production under solid state fermentation conditions (SSF) by using WP as moistening agent instead of distilled water. The optimum conditions for production of B. sphaericus 14N1 on wheat bran-WP medium were 10 g wheat bran/250 ml flask moistened with 10-70% WP at 50% moisture content, inoculum size ranged between 17.2x10(7) and 34.4x10(7) and 6 days incubation under static conditions at 30 degrees C. Preliminary pilot-scale production of B. sphaericus 14N1 under SSF conditions in trays proved that wheat bran-WP medium was efficient and economic for industrial production of mosquitocidal toxin by B. sphaericus.

  10. Product ion filtering with rapid polarity switching for the detection of all fumonisins and AAL-toxins.

    Science.gov (United States)

    Renaud, Justin B; Kelman, Megan J; Qi, Tianyu F; Seifert, Keith A; Sumarah, Mark W

    2015-11-30

    Fumonisins and AAL-toxins are structurally similar mycotoxins that contaminate agricultural crops and foodstuffs. Traditional analytical screening methods are designed to target the known compounds for which standards are available but there is clear evidence that many other derivatives exist and could be toxic. A fast, semi-targeted method for the detection of all known fumonisins, AAL-toxins and related emerging toxins is required. Strains of Fusarium verticillioides, Alternaria arborescens and Aspergillus welwitschiae were grown on their associated crops (maize, tomatoes, and grapes, respectively). Extracts were first analyzed in negative mode using product ion filtering to detect the tricarballylic ester product ion that is common to fumonisins and AAL-toxins (m/z 157.0142). During the same liquid chromatography (LC) run, rapid polarity switching was then used to collect positive mode tandem mass spectrometric (MS(2) ) data for characterization of the detected compounds. Fumonisin B1 , B2 , B3 and B4 were detected on Fusarium contaminated maize, AAL-toxins TA, TB, TD, TE were detected on Alternaria inoculated tomatoes and fumonisin B2 , B4 and B6 on Aspergillus contaminated grapes. Additionally, over 100 structurally related compounds possessing a tricarballylic ester were detected from the mould inoculated plant material. These included a hydroxyl-FB1 from F. verticillioides inoculated maize, keto derivatives of AAL-toxins from A. arborescens inoculated tomatoes, and two previously unreported classes of non-aminated fumonisins from Asp. welwitschiae contaminated grapes. A semi-targeted method for the detection of all fumonisins and AAL-toxins in foodstuffs was developed. The use of the distinctive tricarballylic ester product anion for detection combined with rapid polarity switching and positive mode MS(2) is an effective strategy for differentiating between known isomers such as FB1 and FB6 . This analytical tool is also effective for the identification of

  11. Detection by hyperspectral imaging of shiga toxin-producing Escherichia coli serogroups O26, O45, O103, O111, O121, and O145 on rainbow agar.

    Science.gov (United States)

    Windham, William R; Yoon, Seung-Chul; Ladely, Scott R; Haley, Jennifer A; Heitschmidt, Jerry W; Lawrence, Kurt C; Park, Bosoon; Narrang, Neelam; Cray, William C

    2013-07-01

    The U.S. Department of Agriculture, Food Safety Inspection Service has determined that six non-O157 Shiga toxin-producing Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, and O145) are adulterants in raw beef. Isolate and phenotypic discrimination of non-O157 STEC is problematic due to the lack of suitable agar media. The lack of distinct phenotypic color variation among non-O157serogroups cultured on chromogenic agar poses a challenge in selecting colonies for confirmation. In this study, visible and near-infrared hyperspectral imaging and chemometrics were used to detect and classify non-O157 STEC serogroups grown on Rainbow agar O157. The method was first developed by building spectral libraries for each serogroup obtained from ground-truth regions of interest representing the true identity of each pixel and thus each pure culture colony in the hyperspectral agar-plate image. The spectral library for the pure-culture non-O157 STEC consisted of 2,171 colonies, with spectra derived from 124,347 of pixels. The classification models for each serogroup were developed with a k nearest-neighbor classifier. The overall classification training accuracy at the colony level was 99%. The classifier was validated with ground beef enrichments artificially inoculated with 10, 50, and 100 CFU/ml STEC. The validation ground-truth regions of interest of the STEC target colonies consisted of 606 colonies, with 3,030 pixels of spectra. The overall classification accuracy was 98%. The average specificity of the method was 98% due to the low false-positive rate of 1.2%. The sensitivity ranged from 78 to 100% due to the false-negative rates of 22, 7, and 8% for O145, O45, and O26, respectively. This study showed the potential of visible and near-infrared hyperspectral imaging for detecting and classifying colonies of the six non-O157 STEC serogroups. The technique needs to be validated with bacterial cultures directly extracted from meat products and positive

  12. Detection of Escherichia coli O157:H7 in the beef marketed in Malaysia.

    Science.gov (United States)

    Radu, S; Abdul Mutalib, S; Rusul, G; Ahmad, Z; Morigaki, T; Asai, N; Kim, Y B; Okuda, J; Nishibuchi, M

    1998-03-01

    Twelve strains of Escherichia coli O157:H7 were isolated from 9 of 25 beef samples purchased from retail stores in Malaysia. These strains produced Shiga toxin 2 with or without Shiga toxin 1 and had the eae gene and a 60-MDa plasmid. The antibiograms and the profiles of the arbitrarily primed PCR of the strains were diverse, suggesting that the strains may have originated from diverse sources.

  13. Mexican unpasteurised fresh cheeses are contaminated with Salmonella spp., non-O157 Shiga toxin producing Escherichia coli and potential uropathogenic E. coli strains: A public health risk.

    Science.gov (United States)

    Guzman-Hernandez, Rosa; Contreras-Rodriguez, Araceli; Hernandez-Velez, Rosa; Perez-Martinez, Iza; Lopez-Merino, Ahide; Zaidi, Mussaret B; Estrada-Garcia, Teresa

    2016-11-21

    Fresh cheeses are a main garnish of Mexican food. Consumption of artisanal fresh cheeses is very common and most of them are made from unpasteurised cow milk. A total of 52 fresh unpasteurised cheeses of five different types were purchased from a variety of suppliers from Tabasco, Mexico. Using the most probable number method, 67% and 63% of samples were positive for faecal coliforms and E. coli, respectively; revealing their low microbiological quality. General hygienic conditions and practices of traditional cheese manufacturers were poor; most establishments had unclean cement floors, all lacked windows and doors screens, and none of the food-handlers wore aprons, surgical masks or bouffant caps. After analysing all E. coli isolates (121 strains) for the presence of 26 virulence genes, results showed that 9 (17%) samples were contaminated with diarrheagenic E. coli strains, 8 harboured non-O157 Shiga toxin producing E. coli (STEC), and one sample contained both STEC and diffusely adherent E. coli strains. All STEC strains carried the stx1 gene. Potential uropathogenic E. coli (UPEC) strains were isolated from 15 (29%) samples; the most frequent gene combination was fimA-agn43. Two samples were contaminated with Salmonella. The results demonstrated that unpasteurised fresh cheeses produced in Tabasco are of poor microbiological quality and may frequently harbour foodborne pathogens. Food safety authorities in Mexico need to conduct more rigorous surveillance of fresh cheeses. Furthermore, simple and inexpensive measures as establishing programs emphasizing good hand milking practices and hygienic manufacturing procedures may have a major effect on improving the microbiological quality of these food items. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Deteksi Produksi Toksin Stx-1 dan Stx-2 dari Escherichia coli O157:H7 Isolat Lokal Hasil Isolasi Feses dan Daging Sapi

    Directory of Open Access Journals (Sweden)

    I Wayan Suardana

    2009-12-01

    Full Text Available Shiga toxin produced by Escherichia coli O157:H7 can cause outbreaks and sporadic cases of serioushuman diseases. The diseases are indicated by hemorrhagic colitis and hemolytic uremic syndrome. Meatand meat products have been identified as vehicles of food borne disease caused by E.coli O157:H7. Themain aim of this research was to identify the correlation between the level of E.coli O157:H7 contaminationand the presence of Shiga toxin (Stx1 and Stx2 by applying method of Vero toxin Escherichia coli-ReversePassive Agglutination Test (VTEC-RPLA. The results showed that 3 of 7 isolates and 1 of 4 isolatesisolated from feces of cattle and beef, respectively produced Stx 1 (VT1. In the detection of Stx 2 (VT2, 4of 7 isolates and 1 of 4 isolates, isolated from the same samples were found to produce this toxin.According to all isolates, in this research showed, 1 isolate was found to produce VT2, 4 isolates to produceboth VT1 and VT2, while 6 isolates showed negative results either to VT1 or VT2.

  15. Endoplasmic Reticulum-Targeted Subunit Toxins Provide a New Approach to Rescue Misfolded Mutant Proteins and Revert Cell Models of Genetic Diseases.

    Directory of Open Access Journals (Sweden)

    Humaira Adnan

    Full Text Available Many germ line diseases stem from a relatively minor disturbance in mutant protein endoplasmic reticulum (ER 3D assembly. Chaperones are recruited which, on failure to correct folding, sort the mutant for retrotranslocation and cytosolic proteasomal degradation (ER-associated degradation-ERAD, to initiate/exacerbate deficiency-disease symptoms. Several bacterial (and plant subunit toxins, retrograde transport to the ER after initial cell surface receptor binding/internalization. The A subunit has evolved to mimic a misfolded protein and hijack the ERAD membrane translocon (dislocon, to effect cytosolic access and cytopathology. We show such toxins compete for ERAD to rescue endogenous misfolded proteins. Cholera toxin or verotoxin (Shiga toxin containing genetically inactivated (± an N-terminal polyleucine tail A subunit can, within 2-4 hrs, temporarily increase F508delCFTR protein, the major cystic fibrosis (CF mutant (5-10x, F508delCFTR Golgi maturation (<10x, cell surface expression (20x and chloride transport (2x in F508del CFTR transfected cells and patient-derived F508delCFTR bronchiolar epithelia, without apparent cytopathology. These toxoids also increase glucocerobrosidase (GCC in N370SGCC Gaucher Disease fibroblasts (3x, another ERAD-exacerbated misfiling disease. We identify a new, potentially benign approach to the treatment of certain genetic protein misfolding diseases.

  16. Multiplexed Single Intact Cell Droplet Digital PCR (MuSIC ddPCR) Method for Specific Detection of Enterohemorrhagic E. coli (EHEC) in Food Enrichment Cultures

    OpenAIRE

    McMahon, Tanis C.; Blais, Burton W.; Wong, Alex; Carrillo, Catherine D.

    2017-01-01

    Foodborne illness attributed to enterohemorrhagic E. coli (EHEC), a highly pathogenic subset of Shiga toxin-producing E. coli (STEC), is increasingly recognized as a significant public health issue. Current microbiological methods for identification of EHEC in foods often use PCR-based approaches to screen enrichment broth cultures for characteristic gene markers [i.e., Shiga toxin (stx) and intimin (eae)]. However, false positives arise when complex food matrices, such as beef, contain mixtu...

  17. Shiga toxin-producing Escherichia coli (STEC) O22:H8 isolated from cattle reduces E. coli O157:H7 adherence in vitro and in vivo.

    Science.gov (United States)

    Martorelli, L; Albanese, A; Vilte, D; Cantet, R; Bentancor, A; Zolezzi, G; Chinen, I; Ibarra, C; Rivas, M; Mercado, E C; Cataldi, A

    2017-09-01

    Shiga toxin-producing Escherichia coli (STEC) are a group of bacteria responsible for food-associated diseases. Clinical features include a wide range of symptoms such as diarrhea, hemorrhagic colitis and the hemolytic uremic syndrome (HUS), a life-threatening condition. Our group has observed that animals naturally colonized with STEC strains of unknown serotype were not efficiently colonized with E. coli O157:H7 after experimental infection. In order to assess the basis of the interference, three STEC strains were isolated from STEC persistently-colonized healthy cattle from a dairy farm in Buenos Aires, Argentina. The three isolated strains are E. coli O22:H8 and carry the stx1 and stx2d genes. The activatable activity of Stx2d was demonstrated in vitro. The three strains carry the adhesins iha, ehaA and lpf O113 . E. coli O22:H8 formed stronger biofilms in abiotic surface than E. coli O157:H7 (eae+, stx2+) and displayed a more adherent phenotype in vitro towards HeLa cells. Furthermore, when both serotypes were cultured together O22:H8 could reduce O157:H7 adherence in vitro. When calves were intragastrically pre-challenged with 10 8 CFU of a mixture of the three STEC strains and two days later challenged with the same dose of the strain E. coli O157:H7 438/99, the shedding of the pathogen was significantly reduced. These results suggest that E. coli O22:H8, a serotype rarely associated with human illness, might compete with O157:H7 at the bovine recto-anal junction, making non-O157 carrying-calves less susceptible to O157:H7 colonization and shedding of the bacteria to the environment. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Genomic comparison of Escherichia coli O104:H4 isolates from 2009 and 2011 reveals plasmid, and prophage heterogeneity, including shiga toxin encoding phage stx2.

    Directory of Open Access Journals (Sweden)

    Sanaa A Ahmed

    Full Text Available In May of 2011, an enteroaggregative Escherichia coli O104:H4 strain that had acquired a Shiga toxin 2-converting phage caused a large outbreak of bloody diarrhea in Europe which was notable for its high prevalence of hemolytic uremic syndrome cases. Several studies have described the genomic inventory and phylogenies of strains associated with the outbreak and a collection of historical E. coli O104:H4 isolates using draft genome assemblies. We present the complete, closed genome sequences of an isolate from the 2011 outbreak (2011C-3493 and two isolates from cases of bloody diarrhea that occurred in the Republic of Georgia in 2009 (2009EL-2050 and 2009EL-2071. Comparative genome analysis indicates that, while the Georgian strains are the nearest neighbors to the 2011 outbreak isolates sequenced to date, structural and nucleotide-level differences are evident in the Stx2 phage genomes, the mer/tet antibiotic resistance island, and in the prophage and plasmid profiles of the strains, including a previously undescribed plasmid with homology to the pMT virulence plasmid of Yersinia pestis. In addition, multiphenotype analysis showed that 2009EL-2071 possessed higher resistance to polymyxin and membrane-disrupting agents. Finally, we show evidence by electron microscopy of the presence of a common phage morphotype among the European and Georgian strains and a second phage morphotype among the Georgian strains. The presence of at least two stx2 phage genotypes in host genetic backgrounds that may derive from a recent common ancestor of the 2011 outbreak isolates indicates that the emergence of stx2 phage-containing E. coli O104:H4 strains probably occurred more than once, or that the current outbreak isolates may be the result of a recent transfer of a new stx2 phage element into a pre-existing stx2-positive genetic background.

  19. Protection against Shiga-Toxigenic Escherichia coli by Non-Genetically Modified Organism Receptor Mimic Bacterial Ghosts.

    Science.gov (United States)

    Paton, Adrienne W; Chen, Austen Y; Wang, Hui; McAllister, Lauren J; Höggerl, Florian; Mayr, Ulrike Beate; Shewell, Lucy K; Jennings, Michael P; Morona, Renato; Lubitz, Werner; Paton, James C

    2015-09-01

    Shiga-toxigenic Escherichia coli (STEC) causes severe gastrointestinal infections in humans that may lead to life-threatening systemic sequelae, such as the hemolytic uremic syndrome (HUS). Rapid diagnosis of STEC infection early in the course of disease opens a window of opportunity for therapeutic intervention, for example, by administration of agents that neutralize Shiga toxin (Stx) in the gut lumen. We previously developed a recombinant bacterium that expresses a mimic of the Stx receptor globotriaosyl ceramide (Gb3) on its surface through modification of the lipopolysaccharide (A. W. Paton, R. Morona, and J. C. Paton, Nat Med 6:265-270, 2000, http://dx.doi.org/10.1038/73111). This construct was highly efficacious in vivo, protecting mice from otherwise fatal STEC disease, but the fact that it is a genetically modified organism (GMO) has been a barrier to clinical development. In the present study, we have overcome this issue by development of Gb3 receptor mimic bacterial ghosts (BGs) that are not classified as GMOs. Gb3-BGs neutralized Stx1 and Stx2 in vitro with high efficiency, whereas alternative Gb3-expressing non-GMO subbacterial particles (minicells and outer membrane blebs) were ineffective. Gb3-BGs were highly efficacious in a murine model of STEC disease. All mice (10/10) treated with Gb3-BGs survived challenge with a highly virulent O113:H21 STEC strain and showed no pathological signs of renal injury. In contrast, 6/10 mice treated with control BGs succumbed to STEC challenge, and survivors exhibited significant weight loss, neutrophilia, and histopathological evidence of renal damage. Thus, Gb3-BGs offer a non-GMO approach to treatment of STEC infection in humans, particularly in an outbreak setting. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  20. Postdiarrheal Shiga Toxin-Mediated Hemolytic Uremic Syndrome Similar to Septic Shock Síndrome urémico hemolítico símil shock séptico, posterior a diarrea mediada por toxina shiga

    Directory of Open Access Journals (Sweden)

    Patricia G. Valles

    2005-10-01

    Full Text Available The inflammatory response of host endothelial cells is included in the development of vascular damage observed in enterohemorrhagic Escherichia coli (EHEC infection, resulting in hemolytic uremic syndrome (HUS. The response to a non-conventional treatment for a group of D+ HUS (diarrhea positive HUS patients, with clinical hemodynamic parameters of septic shock was evaluated in this prospective study (1999-2003. Twelve children 2.8 ± 0.6 years old, with D+ HUS produced by E. coli infection with serological evidence of Shiga toxin, presenting severe unstable hemodynamic parameters and neurological dysfunction at onset, were studied. The protocol included fresh frozen plasma infusions, methylprednisolone pulses (10mg/k/day for three consecutive days and plasma exchange for five days, starting after admission to the intensive care unit (ICU. The twelve patients with increased pediatric risk of mortality (PRISM score: 18 ± 2 after admission to intensive care unit (ICU, required dialysis for 17.4 ± 4 days, mechanical ventilator assistance for 10 ± 1 days and early inotropic drugs support for 10.5 ± 1 days. Neurological dysfunction included generalized tonic-clonic seizures lasting for 5.4 ± 1 days, n:8. Focal seizures were present in the remaining patients. Dilated cardiomyopathy was present in 6 children. Eight children suffered hemorrhagic colitis. Nine patients survived. Within one year of the injury, neurological sequelae, Glasgow outcome scale (GOS 3 and 4, were present in two patients, chronic renal failure in one patient. We suggest that early introduction of this protocol could benefit D+ HUS patients with hemodynamic instability and neurological dysfunction at onset. Further studies are likely to elucidate the mechanisms involved in this early adverse clinical presentation of D+ HUS patients.La respuesta inflamatoria de la célula endotelial se incluye en el desarrollo del daño vascular observado en la infección por Escherichia coli

  1. Comparison of toxin production by clostridium botulinum type E in irradiated vacuum-packed trout (Salmo gairdneri)

    International Nuclear Information System (INIS)

    Hussain, A.M.; Ehlermann, D.; Diehl, J.F.

    1977-01-01

    Trouts obtained from a nearby Fish farm were slaughtered, gutted, cut into 100 g samples and inoculated with 10 1 , 10 3 and 10 5 spores per g of Clostridium botulinum type E. The vacuum-packed samples were stored under melting ice (0 0 C) and at temperatures of 5 0 and 10 0 C for periods of up to 8 weeks. At weekly intervals, occurrence of spoilage and toxin production were determined. Only at 10 0 C storage, the irradiated samples showed toxin production before spoilage was observed. When the fishes were stored at 5 0 C, no toxicity occurred before spoilage was observed even in samples treated with doses as high as 200 krad. Samples stored under melting ice, irradiated or unirradiated, never showed toxin production. It is concluded that the radurization of fish at doses of about 100 or 200 krad and at storage temperatures of melting ice or up to 5 0 C is safe with regard to a possible botulism risk. (orig.) [de

  2. Survey of environmental radiation dose rates in Kyoto and Shiga prefectures, Japan

    International Nuclear Information System (INIS)

    Minamia, Kazuyuki; Shimo, Michikuni; Oka, Mitsuaki; Ejiri, Kazutaka; Sugino, Masato; Minato, Susumu; Hosoda, Masahiro; Yamada, Junya; Fukushi, Masahiro

    2008-01-01

    We have measured environmental radiation dose rates in several Prefectures, such as Ai chi Prefecture, Gifu Prefecture, and Mie Prefecture, in central Japan. Recently, we measured the environmental radiation dose rates in Kyoto and Shiga Prefectures that are also located in central Japan with a car-borne survey system. At the time of measurement, Kyoto Prefecture (area: 4,613 km 2 ) had a total of 36 districts, and Shiga Prefecture (area: 3,387 km 2 ) a total of 26. Terrestrial gamma ray dose rates and secondary cosmic ray dose rates were measured by a 2 inches ψ x 2 inches NaI(Tl) scintillation counter and a handy-type altimeter (GPS eTrex Legend by Gamin), respectively. The following factors were taken into consideration the shielding effect of the car body, the effect of the road pavement, radon progeny borne by precipitation, and increases in tunnels and near the walls. Terrestrial gamma ray dose rates in Kyoto and Shiga Prefectures were estimated to be 51.7 ± 6.0 n Gy/h (district average: 52.4 ± 4.7 n Gy/h), 52.2 ± 10.5 n Gy/h (district average: 51.9 ± 8.1 n Gy/h), respectively. Secondary cosmic ray dose rates in Kyoto and Shiga Prefectures were 30.0 ± 0.6 n Gy/h (district average: 29.9 ±0.3 n Gy/h), 30.1 ± 0.3 n Gy/h (district average: 30.0 ± 0.2 n Gy/h), respectively. The environmental radiation dose rates due to the sum dose rates of terrestrial gamma ray and secondary cosmic ray in Kyoto and Shiga Prefectures were 81.7 ± 6.2 n Gy/h (district average: 82.3 ± 4.8 n Gy/h), 82.3 ± 10.6 n Gy/h (district average: 82.0 ± 8.1 n Gy/h), respectively. We confirmed that the environmental radiation dose rates in Kyoto and Shiga Prefectures mainly depended on the change of the terrestrial gamma ray dose rates, since the secondary cosmic ray dose rates had little change. Therefore, radiation dose-rate maps of the terrestrial gamma rays as well as maps of the environmental radiation dose-rate were drawn. (author)

  3. Occurrence and characterization of Shiga toxin-producing Escherichia coli O157:H7 and other non-sorbitol-fermenting E. coli in cattle and humans in urban areas of Morogoro, Tanzania.

    Science.gov (United States)

    Lupindu, Athumani M; Olsen, John E; Ngowi, Helena A; Msoffe, Peter L M; Mtambo, Madundo M; Scheutz, Flemming; Dalsgaard, Anders

    2014-07-01

    Escherichia coli strains such as Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli, enterotoxigenic, attaching, and effacing E. coli, and enteroinvasive E. coli cause diarrhea in humans. Although other serotypes exist, the most commonly reported STEC in outbreaks is O157:H7. A cross-sectional study was conducted to isolate and characterize non-sorbitol-fermenting (NSF) E. coli O157:H7 from urban and periurban livestock settings of Morogoro, Tanzania. Human stool, cattle feces, and soil and water samples were collected. Observations and questionnaire interview studies were used to gather information about cattle and manure management practices in the study area. E. coli were isolated on sorbitol MacConkey agar and characterized by conventional biochemical tests. Out of 1049 samples, 143 (13.7%) yielded NSF E. coli. Serological and antimicrobial tests and molecular typing were performed to NSF E. coli isolates. These procedures detected 10 (7%) pathogenic E. coli including STEC (n=7), enteropathogenic E. coli (EPEC) (n=2), and attaching and effacing E. coli (A/EEC) (n=1) strains. The STEC strains had the ability to produce VT1 and different VT2 toxin subtypes that caused cytopathic effects on Vero cells. The prevalence of STEC in cattle was 1.6%, out of which 0.9% was serotype O157:H7 and the overall prevalence of diarrheagenic E. coli in cattle was 2.2%. The serotypes O157:H7, O142:H34, O113:H21, O+:H-, O+:H16, and O25:H4 were identified. One ESBL-producing isolate showed the MLST type ST131. To our knowledge, this is the first finding in Tanzania of this recently emerged worldwide pandemic clonal group, causing widespread antimicrobial-resistant infections, and adds knowledge of the geographical distribution of ST131. Cattle manure was indiscriminately deposited within residential areas, and there was direct contact between humans and cattle feces during manure handling. Cattle and manure management practices expose humans, animals, and the environment

  4. The Trojan Horse of the microbiological arms race: phage-encoded toxins as a defence against eukaryotic predators.

    Science.gov (United States)

    Arnold, Jason W; Koudelka, Gerald B

    2014-02-01

    Phage-encoded Shiga toxin (Stx) acts as a bacterial defence against the eukaryotic predator Tetrahymena. To function as an effective bacterial anti-predator defence, Stx must kill a broad spectrum of predators. Consistent with that assertion, we show here that bacterially encoded Stx efficiently kills the bacteriovore Acanthamoeba castellanii in co-culture. We also show that, in addition to Stx, the phage-encoded exotoxin, diphtheria toxin (Dtx) expressed by Corynebacterium diphtheriae also can function as part of an anti-predator strategy; it kills Acanthamoeba in co-culture. Interestingly, only exotoxins produced by bacteria internalized by the Acanthamoeba predator are cytolethal; the presence of purified Dtx or Stx in culture medium has no effect on predator viability. This finding is consistent with our results indicating that intoxication of Acanthamoeba by these exotoxins does not require a receptor. Thus bacteria, in the disguise of a food source, function as a 'Trojan Horse', carrying genes encoding an exotoxin into target organisms. This 'Trojan Horse' mechanism of exotoxin delivery into predator cells allows intoxication of predators that lack a cell surface receptor for the particular toxin, allowing bacteria-bearing exotoxins to kill a broader spectrum of predators, increasing the fitness of the otherwise 'defenceless' prey bacteria. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

  5. Engineering toxins for 21st century therapies.

    Science.gov (United States)

    Chaddock, John A; Acharya, K Ravi

    2011-04-01

    'Engineering Toxins for 21st Century Therapies' (9-10 September 2010) was part of the Royal Society International Seminar series held at the Kavli International Centre, UK. Participants were assembled from a range of disciplines (academic, industry, regulatory, public health) to discuss the future potential of toxin-based therapies. The meeting explored how the current structural and mechanistic knowledge of toxins could be used to engineer future toxin-based therapies. To date, significant progress has been made in the design of novel recombinant biologics based on domains of natural toxins, engineered to exhibit advantageous properties. The meeting concluded, firstly that future product development vitally required the appropriate combination of creativity and innovation that can come from the academic, biotechnology and pharma sectors. Second, that continued investigation into understanding the basic science of the toxins and their targets was essential in order to develop new opportunities for the existing products and to create new products with enhanced properties. Finally, it was concluded that the clinical potential for development of novel biologics based on toxin domains was evident. © 2011 The Authors Journal compilation © 2011 FEBS.

  6. Dysport: pharmacological properties and factors that influence toxin action.

    Science.gov (United States)

    Pickett, Andy

    2009-10-01

    The pharmacological properties of Dysport that influence toxin action are reviewed and compared with other botulinum toxin products. In particular, the subject of diffusion is examined and discussed based upon the evidence that currently exists, both from laboratory studies and from clinical data. Diffusion of botulinum toxin products is not related to the size of the toxin complex in the product since the complex dissociates under physiological conditions, releasing the naked neurotoxin to act. The active neurotoxin in Type A products is the same and therefore diffusion is equal when equal doses are administered.

  7. Timeliness of Surveillance during Outbreak of Shiga Toxin–producing Escherichia coli Infection, Germany, 2011

    OpenAIRE

    Altmann, Mathias; Wadl, Maria; Altmann, Doris; Benzler, Justus; Eckmanns, Tim; Krause, Gérard; Spode, Anke; an der Heiden, Matthias

    2011-01-01

    In the context of a large outbreak of Shiga toxin–producing Escherichia coli O104:H4 in Germany, we quantified the timeliness of the German surveillance system for hemolytic uremic syndrome and Shiga toxin–producing E. coli notifiable diseases during 2003–2011. Although reporting occurred faster than required by law, potential for improvement exists at all levels of the information chain.

  8. Timeliness of Surveillance during Outbreak of Shiga Toxin–producing Escherichia coli Infection, Germany, 2011

    Science.gov (United States)

    Wadl, Maria; Altmann, Doris; Benzler, Justus; Eckmanns, Tim; Krause, Gérard; Spode, Anke; an der Heiden, Matthias

    2011-01-01

    In the context of a large outbreak of Shiga toxin–producing Escherichia coli O104:H4 in Germany, we quantified the timeliness of the German surveillance system for hemolytic uremic syndrome and Shiga toxin–producing E. coli notifiable diseases during 2003–2011. Although reporting occurred faster than required by law, potential for improvement exists at all levels of the information chain. PMID:22000368

  9. The effect of irradiation on the viability and toxin production of various species of fungi

    International Nuclear Information System (INIS)

    Sellyei, Gy-ne.; Vanyi, A.; Petri, A.

    1979-01-01

    The procedure to sterilize various kinds of food by irradiation has been used successfully for many years. The irradiation resistance of fodder contaminating moulds and their toxins was examined. Irradiation, heat treatment, and irradiation combined with heat treatment were used in experiments with Fusarium. However, only irradiation was used with the conidium suspensions of Aspergillus and Penicillium. After an appropriate time of incubation, the survival and the toxin producing capacity were determined. The heat treatment killed Fusarium but failed to kill Aspergillus and Penicillium. Owing to irradiation the reproductive capacity all of the fungi tested decreased, but in the surviving cultures the mycotoxin production increased. Using heat treatment combined with irradiation the toxin content of samples reached a high level in a short time and became stabilized. (author)

  10. Enterohemorrhagic Escherichia coli Hybrid Pathotype O80:H2 as a New Therapeutic Challenge

    Science.gov (United States)

    Soysal, Nurcan; Mariani-Kurkdjian, Patricia; Smail, Yasmine; Liguori, Sandrine; Gouali, Malika; Loukiadis, Estelle; Fach, Patrick; Bruyand, Mathias; Blanco, Jorge; Bidet, Philippe

    2016-01-01

    We describe the epidemiology, clinical features, and molecular characterization of enterohemorrhagic Escherichia coli (EHEC) infections caused by the singular hybrid pathotype O80:H2, and we examine the influence of antibiotics on Shiga toxin production. In France, during 2005–2014, a total of 54 patients were infected with EHEC O80:H2; 91% had hemolytic uremic syndrome. Two patients had invasive infections, and 2 died. All strains carried stx2 (variants stx2a, 2c, or 2d); the rare intimin gene (eae-ξ); and at least 4 genes characteristic of pS88, a plasmid associated with extraintestinal virulence. Similar strains were found in Spain. All isolates belonged to the same clonal group. At subinhibitory concentrations, azithromycin decreased Shiga toxin production significantly, ciprofloxacin increased it substantially, and ceftriaxone had no major effect. Antibiotic combinations that included azithromycin also were tested. EHEC O80:H2, which can induce hemolytic uremic syndrome complicated by bacteremia, is emerging in France. However, azithromycin might effectively combat these infections. PMID:27533474

  11. A review of current knowledge on toxic benthic freshwater cyanobacteria--ecology, toxin production and risk management.

    Science.gov (United States)

    Catherine, Quiblier; Susanna, Wood; Isidora, Echenique-Subiabre; Mark, Heath; Aurélie, Villeneuve; Jean-François, Humbert

    2013-10-01

    Benthic cyanobacteria are found globally in plethora of environments. Although they have received less attention than their planktonic freshwater counterparts, it is now well established that they produce toxins and reports of their involvement in animal poisonings have increased markedly during the last decade. Most of the known cyanotoxins have been identified from benthic cyanobacteria including: the hepatotoxic microcystins, nodularins and cylindrospermopsins, the neurotoxic saxitoxins, anatoxin-a and homoanatoxin-a and dermatotoxins, such as lyngbyatoxin. In most countries, observations of toxic benthic cyanobacteria are fragmented, descriptive and in response to animal toxicosis events. Only a limited number of long-term studies have aimed to understand why benthic proliferations occur, and/or how toxin production is regulated. These studies have shown that benthic cyanobacterial blooms are commonly a mixture of toxic and non-toxic genotypes and that toxin concentrations can be highly variable spatially and temporally. Physiochemical parameters responsible for benthic proliferation vary among habitat type with physical disturbance (e.g., flow regimes, wave action) and nutrients commonly identified as important. As climatic conditions change and anthropogenic pressures on waterways increase, it seems likely that the prevalence of blooms of benthic cyanobacteria will increase. In this article we review current knowledge on benthic cyanobacteria: ecology, toxin-producing species, variables that regulate toxin production and bloom formation, their impact on aquatic and terrestrial organisms and current monitoring and management strategies. We suggest research needs that will assist in filling knowledge gaps and ultimately allow more robust monitoring and management protocols to be developed. Copyright © 2013 Elsevier Ltd. All rights reserved.

  12. Actualización en el tratamiento del síndrome urémico hemolítico endémico: Patogénesis y tratamiento de la complicación sistémica más grave de las infecciones por Escherichia coli productor de toxina Shiga Update on the treatment of endemic hemolytic uremic syndrome: Pathogenesis and treatment of the most severe systemic complication of infections by Shiga toxin-producing Escherichia coli

    Directory of Open Access Journals (Sweden)

    Romina J. Fernández-Brando

    2011-08-01

    Full Text Available La forma típica o post-diarreica del síndrome urémico hemolítico (SUH es la complicación más grave de las infecciones por cepas de Escherichia coli productoras de toxina Shiga (STEC. En la Argentina el SUH es un problema crítico de salud pública, ya que representa la principal causa de falla renal aguda en la infancia, la segunda causa de falla renal crónica, y aporta el 20% de los casos de transplante renal durante la infancia y la adolescencia. A pesar de los avances en el conocimiento de su patogénesis, el único tratamiento actual de los pacientes con SUH es de sostén, y no existen terapias específicas ni preventivas. En la presente revisión expondremos los conocimientos básicos de los mecanismos patogénicos y discutiremos los enfoques terapéuticos tradicionales e innovadores, con especial foco en la situación nacional y los aportes hechos por grupos de la Argentina.The typical form of hemolytic uremic syndrome (HUS is the major complication of Shiga toxin-producing Escherichia coli (STEC infections. HUS is a critical health problem in Argentina since it is the main cause of acute renal failure in children and the second cause of chronic renal failure, giving account for 20% of renal transplants in children and adolescents in our country. In spite of the extensive research in the field, the mainstay of treatment for patients with HUS is supportive therapy, and there are no specific therapies preventing or ameliorating the disease course. In this review, we present the current knowledge about pathogenic mechanisms and discuss traditional and innovative therapeutic approaches, with special focus in national status and contributions made by Argentinean groups.

  13. [Production of pertussis toxin by Bordetella pertussis strains isolated from patients with whooping cough].

    Science.gov (United States)

    Zaĭtsev, E M; Mertsalova, N U; Shinkarev, A S; Mazurova, I K; Zakharova, N S

    2011-01-01

    To assess level of pertussin toxin (PT) production by vaccine strains of Bordetella pertussis and strains isolated from patients with whooping cough. Concentration of PT in supernatants of microbial cultures of 3 vaccine strains and 25 strains of B. pertussis isolated from patients with pertussis in 2001 - 2005 was measured with enzyme immunoassay using gamma-globulin fractions of rabbit antiserum to PT as immunosorbent or included in peroxidase conjugates. Level of PT production by strains isolated from infected persons varied from 3 +/- 0.5 to 64.8 +/- 12.2 ng/MFU/ml: in 9 strains--from 3 +/- 0.5 to 9.4 +/- 2.1 ng/MFU/ml, in 7--10.5 +/- 1.8 to 18.4 +/- 2.6 ng/MFU/ml, and in 9--23.6 +/- 4.5 to 64.8 +/- 12.2 ng/MFU/ml. B. pertussis strains isolated from patients were heterogeneous on level of PT production. Difference in expression of PT between strains were as high as 20-fold. Conditionally low, moderate and high levels of PT production had 9 (36%), 7 (28%), and 9 (36%) of 25 studied strains. Three vaccine strains had levels of toxin production similar to recently isolated strains with moderate level of its production.

  14. A novel mechanism of bacterial toxin transfer within host blood cell-derived microvesicles.

    Directory of Open Access Journals (Sweden)

    Anne-lie Ståhl

    2015-02-01

    Full Text Available Shiga toxin (Stx is the main virulence factor of enterohemorrhagic Escherichia coli, which are non-invasive strains that can lead to hemolytic uremic syndrome (HUS, associated with renal failure and death. Although bacteremia does not occur, bacterial virulence factors gain access to the circulation and are thereafter presumed to cause target organ damage. Stx was previously shown to circulate bound to blood cells but the mechanism by which it would potentially transfer to target organ cells has not been elucidated. Here we show that blood cell-derived microvesicles, shed during HUS, contain Stx and are found within patient renal cortical cells. The finding was reproduced in mice infected with Stx-producing Escherichia coli exhibiting Stx-containing blood cell-derived microvesicles in the circulation that reached the kidney where they were transferred into glomerular and peritubular capillary endothelial cells and further through their basement membranes followed by podocytes and tubular epithelial cells, respectively. In vitro studies demonstrated that blood cell-derived microvesicles containing Stx undergo endocytosis in glomerular endothelial cells leading to cell death secondary to inhibited protein synthesis. This study demonstrates a novel virulence mechanism whereby bacterial toxin is transferred within host blood cell-derived microvesicles in which it may evade the host immune system.

  15. Effect of initial O2 and CO2 and low-dose irradiation on toxin production by Clostridium botulinum in MAP fresh pork

    International Nuclear Information System (INIS)

    Lambert, A.D.; Smith, J.P.; Dodds, K.L.

    1991-01-01

    The effects of irradiation, initial O2, initial CO2 and the presence of an O2 and CO2 absorbent on toxin production by Clostridium botulinum in inoculated pork stored at 15 degrees C were studied using a factorial experiment. Toxin production occurred faster in samples initially packaged with 20% O2, compared to samples packaged with 100% N2. The presence of CO2 in the package headspace was not a significant factor affecting time until toxin detection. Irradiation was significant in delaying the time until toxin detection in samples initially packaged with 20% O2 but not in other treatments. Sensory rejection, based primarily on discoloration, occurred within 7 to 14 d, irrespective of treatment. All samples were spoiled before they became toxic

  16. Variability of antibiotic susceptibility and toxin production of Staphylococcus aureus strains isolated from skin, soft tissue, and bone related infections

    NARCIS (Netherlands)

    Sina, Haziz; Ahoyo, Theodora A.; Moussaoui, Wardi; Keller, Daniel; Bankole, Honore S.; Barogui, Yves; Stienstra, Ymkje; Kotchoni, Simeon O.; Prevost, Gilles; Baba-Moussa, Lamine

    2013-01-01

    Background: Staphylococcus aureus is an opportunistic commensal bacterium that mostly colonizes the skin and soft tissues. The pathogenicity of S. aureus is due to both its ability to resist antibiotics, and the production of toxins. Here, we characterize a group of genes responsible for toxin

  17. Diffusion of Botulinum Toxins

    Directory of Open Access Journals (Sweden)

    Matthew A. Brodsky

    2012-08-01

    Full Text Available Background: It is generally agreed that diffusion of botulinum toxin occurs, but the extent of the spread and its clinical importance are disputed. Many factors have been suggested to play a role but which have the most clinical relevance is a subject of much discussion.Methods: This review discusses the variables affecting diffusion, including protein composition and molecular size as well as injection factors (e.g., volume, dose, injection method. It also discusses data on diffusion from comparative studies in animal models and human clinical trials that illustrate differences between the available botulinum toxin products (onabotulinumtoxinA, abobotulinumtoxinA, incobotulinumtoxinA, and rimabotulinumtoxinB.Results: Neither molecular weight nor the presence of complexing proteins appears to affect diffusion; however, injection volume, concentration, and dose all play roles and are modifiable. Both animal and human studies show that botulinum toxin products are not interchangeable, and that some products are associated with greater diffusion and higher rates of diffusion-related adverse events than others.Discussion: Each of the botulinum toxins is a unique pharmacologic entity. A working knowledge of the different serotypes is essential to avoid unwanted diffusion-related adverse events. In addition, clinicians should be aware that the factors influencing diffusion may range from properties intrinsic to the drug to accurate muscle selection as well as dilution, volume, and dose injected.

  18. Imported edible leaves collected at retail sale in England during 2017 with an emphasis on betel and curry leaves: Microbiological quality with respect to Salmonella, Shiga-toxin-producing E. coli (STEC) and levels of Escherichia coli.

    Science.gov (United States)

    McLauchlin, J; Aird, H; Charlett, A; Chattaway, M; Elviss, N; Hartman, H; Jenkins, C; Jørgensen, F; Larkin, L; Sadler-Reeves, L; Willis, C

    2018-05-26

    to investigate the microbiological quality of imported fresh leaves on retail-sale during 2017 with respect to Salmonella, Shiga-toxin-producing Escherichia coli (STEC) and levels of E.coli. 279 samples of imported edible leaves (69 banana, 77 betel, 118 curry and 15 other types) were tested. Salmonella spp. which were confirmed by whole genome sequencing and isolated from 44 samples, 26% from curry leaves, 14% from betel and 2.4% from all other leaf types: 80% of all samples contained ≥10 2 , 44% ≥10 3 and 22% ≥10 4 cfu of E.coli cfu/g. All samples where Salmonella were detected also yielded ≥20 cfu of E.coli/g. 54 samples were tested for STEC which was detected in 6 samples and isolated from three: one was identified as STEC O157:H7. this report further highlights an ongoing problem of Salmonella contamination of imported fresh edible leaves. amongst all food tested by Public Health England (approximately 11,000 per annum), curry leaves were the herb most commonly contaminated with Salmonella, and betel leaves were the most commonly contaminated ready-to-eat food. The high proportion with unsatisfactory E. coli levels and the detection of STEC suggests risks of contamination by multiple enteric pathogens. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  19. Comparison of toxin production by clostridium botulinum type E in irradiated and unirradiated vacuum-packed trout (Salmo gairdneri)

    International Nuclear Information System (INIS)

    Hussain, M.; Ehlermann, D.; Diehl, J.F.

    1977-01-01

    Trouts obtained from a nearby Fish farm were slaughtered, gutted, cut into 100g samples and inoculated with 10 1 , 10 3 and 10 5 spores per g of Clostridium botulinum type E. The vacuum-packed samples were stored under melting ice (0 0 C) and at temperatures of 5 0 and 10 0 C for periods of up to 8 weeks. At weekly intervals, occurrence of spoilage and toxin production were determined. Only at 10 0 C storage, the irradiated samples showed toxin production before spoilage was observed. When the fishes were stored at 5 0 C, no toxicity occured before spoilage was observed even in samples treated with doses as high as 200 krad. Samples stored under melting ice, irradiated or unirradiated, never showed toxin production. It is concluded that the radurization of fish at doses of about 100 or 200 krad and at storage temperatures of melting ice or up to 5 0 C is safe with regard to a possible botulism risk. (orig.) [de

  20. Comparison of toxin production by clostridium botulinum type E in irradiated and unirradiated vacuum-packed trout (Salmo gairdneri)

    International Nuclear Information System (INIS)

    Hussain, A.M.; Ehlermann, D.; Diehl, J.F.

    1977-01-01

    Trouts obtained from a nearby fish farm were slaughtered, gutted, cut into 100 g samples and inoculated with 10 1 , 10 3 and 10 5 spores per g of Clostridium botulinum type E. The vacuum-packed samples were stored under melting ice (0 0 C) and at temperatures of 5 0 and 10 0 C for periods of up to 8 weeks. At weekly intervals, occurrence of spoilage and toxin production were determined. Only at 10 0 C storage, the irradiated samples showed toxin production before spoilage was observed. When the fishes were stored at 5 0 C, no toxicity occurred before spoilage was observed even in samples treated with doses as high as 200 krad. Samples stored under melting ice, irradiated or unirradiated, never showed toxin production. It is concluded that the radurization of fish at doses of about 100 or 200 krad and at storage temperatures of melting ice or up to 5 0 C is safe with regard to a possible botulism risk. (orig.) [de

  1. Effect of Equilibrated pH and Indigenous Spoilage Microorganisms on the Inhibition of Proteolytic Clostridium botulinum Toxin Production in Experimental Meals under Temperature Abuse.

    Science.gov (United States)

    Golden, Max C; Wanless, Brandon J; David, Jairus R D; Lineback, D Scott; Talley, Ryan J; Kottapalli, Bala; Glass, Kathleen A

    2017-08-01

    Clostridium botulinum is a foreseeable biological hazard in prepared refrigerated meals that needs to be addressed in food safety plans. The objective of this study was to evaluate the effect of product composition and storage temperature on the inhibition of botulinum toxin formation in nine experimental meals (meat, vegetable, or carbohydrate based). Treatments were inoculated with proteolytic C. botulinum, vacuum packaged, cooked at 90°C for 10 min, and assayed for botulinum toxin in samples stored at 25°C for up to 96 h for phase 1, or at 25°C for 12 h and then transferred to 12.5°C for up to 12 and 6 weeks in phases 1 and 2, respectively. For phase 1, none of the treatments (equilibrated pH 5.8) supported toxin production when stored at 25°C for 48 h, but toxin production was observed in all treatments at 72 h. For the remaining experiments with storage at 12.5°C, toxin production was dependent on equilibrated pH, storage time, and growth of indigenous spoilage microorganisms. In phase 1, no gross spoilage and no botulinum toxin was detected for any treatment (pH ≤5.8) stored at 12.5°C for 12 weeks. In phase 2, gross spoilage varied by commodity, with the brussels sprouts meal with pH 6.5 showing the most rapid spoilage within 2 weeks and botulinum toxin detected at 5 and 6 weeks for the control and cultured celery juice treatments, respectively. In contrast, spoilage microbes decreased the pH of a pH 5.9 beef treatment by 1.0 unit, potentially inhibiting C. botulinum through 6 weeks at 12.5°C. None of the other treatments with pH 5.8 or below supported toxin production or spoilage. This study provides validation for preventive controls in refrigerated meals. These include equilibrated product pH and storage temperature and time to inhibit toxin formation by proteolytic C. botulinum, but the impact of indigenous microflora on safety and interpretation of challenge studies is also highlighted.

  2. Small-Molecule Inhibitor Leads of Ribosome-Inactivating Proteins Developed Using the Doorstop Approach

    Science.gov (United States)

    2011-03-01

    Medical Systems, Fort Detrick, Maryland, United States of America Introduction Shiga toxin (Stx) produced by the bacteria Shigella dysenteriae and Shiga...the exception that Vero cells (ATCC CCL-81) (green African monkey ) replaced Sp2/0-Ag14 [73]. Vero cells were maintained in EMEM medium (Eagle’s minimum

  3. Recombinant Alpha, Beta, and Epsilon Toxins of Clostridium perfringens: Production Strategies and Applications as Veterinary Vaccines

    Directory of Open Access Journals (Sweden)

    Marcos Roberto A. Ferreira

    2016-11-01

    Full Text Available Clostridium perfringens is a spore-forming, commensal, ubiquitous bacterium that is present in the gastrointestinal tract of healthy humans and animals. This bacterium produces up to 18 toxins. The species is classified into five toxinotypes (A–E according to the toxins that the bacterium produces: alpha, beta, epsilon, or iota. Each of these toxinotypes is associated with myriad different, frequently fatal, illnesses that affect a range of farm animals and humans. Alpha, beta, and epsilon toxins are the main causes of disease. Vaccinations that generate neutralizing antibodies are the most common prophylactic measures that are currently in use. These vaccines consist of toxoids that are obtained from C. perfringens cultures. Recombinant vaccines offer several advantages over conventional toxoids, especially in terms of the production process. As such, they are steadily gaining ground as a promising vaccination solution. This review discusses the main strategies that are currently used to produce recombinant vaccines containing alpha, beta, and epsilon toxins of C. perfringens, as well as the potential application of these molecules as vaccines for mammalian livestock animals.

  4. Climate change impacts on natural toxins in food production systems, exemplified by deoxynivalenol in wheat and diarrhetic shellfish toxins.

    Science.gov (United States)

    van der Fels-Klerx, H J; Olesen, J E; Naustvoll, L-J; Friocourt, Y; Mengelers, M J B; Christensen, J H

    2012-01-01

    Climate change is expected to affect food and feed safety, including the occurrence of natural toxins in primary crop and seafood production; however, to date, quantitative estimates are scarce. This study aimed to estimate the impact of climate change effects on mycotoxin contamination of cereal grains cultivated in the terrestrial area of north west Europe, and on the frequency of harmful algal blooms and contamination of shellfish with marine biotoxins in the North Sea coastal zone. The study focused on contamination of wheat with deoxynivalenol, and on abundance of Dinophysis spp. and the possible relationship with diarrhetic shellfish toxins. The study used currently available data and models. Global and regional climate models were combined with models of crop phenology, mycotoxin prediction models, hydrodynamic models and ecological models, with the output of one model being used as input for the other. In addition, statistical data analyses using existing national datasets from the study area were performed to obtain information on the relationships between Dinophysis spp. cell counts and contamination of shellfish with diarrhetic shellfish toxins as well as on frequency of cereal cropping. In this paper, a summary of the study is presented, and overall conclusions and recommendations are given. Climate change projections for the years 2031-2050 were used as the starting point of the analyses relative to a preceding 20-year baseline period from which the climate change signal was calculated. Results showed that, in general, climate change effects lead to advanced flowering and harvest of wheat, and increased risk of contamination of wheat with deoxynivalenol. Blooms of dinoflagellates were estimated to occur more often. If the group of Dinophysis spp. behaves similarly to other flagellates in the future then frequency of harmful algal blooms of Dinophysis spp. may also increase, but consequences for contamination of shellfish with diarrhetic shellfish

  5. Thermal Responses of Growth and Toxin Production in Four Prorocentrum Species from the Central Red Sea

    KAUST Repository

    Aynousah, Arwa

    2017-06-01

    Harmful algae studies, in particular toxic dinoflagellates, and their response to global warming in the Red Sea are still limited. This study was aimed to be the first to characterize the identity, thermal responses and toxin production of four Prorocentrum strains isolated from the Central Red Sea, Saudi Arabia. Morphological and molecular phylogenetic analysis identified the strains as P. elegans, P. rhathymum and P. emarginatum. However, the identity of strain P. sp.6 is currently unresolved, albeit sharing close affinity with P. leve. Growth experiments showed that all species could grow at 24-32°C, but only P. sp.6 survived the 34°C treatment. The optimum temperatures (Topt) estimated from the Gaussian model corresponded to 27.17, 29.33, 26.87, and 27.64°C for P. sp.6, P. elegans, P. rhathymum and P. emarginatum, respectively. However, some discrepancy with the Topt derived from the growth performance were observed for P. elegans and P. emarginatum, as thermal responses differed from the typical Gaussian fit. The Prorocentrum species examined showed a sharp decrease after the optimum temperature resulting in very high activation energies for the fall slope, especially for P. elegans and P. emarginatum. The minimum critical temperature limit for growth was not detected within the range of temperatures examined. Subsequently, high performance liquid chromatography coupled with mass spectrometry (HPLC-MS) analysis revealed all species as non okadaic acid (OA, common toxin of the Prorocentrum genus) producers at any temperature treatment. However, other forms of toxin (i.e. fast acting toxins) not examined here could be produced. Therefore, further investigations are required. The results of this study provided significant contribution to our knowledge regarding the presence, thermal response and toxin production of four Prorocentrum species from the Central Red Sea, Saudi Arabia.

  6. Military Importance of Natural Toxins and Their Analogs

    Directory of Open Access Journals (Sweden)

    Vladimír Pitschmann

    2016-04-01

    Full Text Available Toxin weapon research, development, production and the ban on its uses is an integral part of international law, with particular attention paid to the protection against these weapons. In spite of this, hazards associated with toxins cannot be completely excluded. Some of these hazards are also pointed out in the present review. The article deals with the characteristics and properties of natural toxins and synthetic analogs potentially constituting the basis of toxin weapons. It briefly describes the history of military research and the use of toxins from distant history up to the present age. With respect to effective disarmament conventions, it mentions certain contemporary concepts of possible toxin applications for military purposes and the protection of public order (suppression of riots; it also briefly refers to the question of terrorism. In addition, it deals with certain traditional as well as modern technologies of the research, synthesis, and use of toxins, which can affect the continuing development of toxin weapons. These are, for example, cases of new toxins from natural sources, their chemical synthesis, production of synthetic analogs, the possibility of using methods of genetic engineering and modern biotechnologies or the possible applications of nanotechnology and certain pharmaceutical methods for the effective transfer of toxins into the organism. The authors evaluate the military importance of toxins based on their comparison with traditional chemical warfare agents. They appeal to the ethics of the scientific work as a principal condition for the prevention of toxin abuse in wars, military conflicts, as well as in non-military attacks.

  7. Military Importance of Natural Toxins and Their Analogs.

    Science.gov (United States)

    Pitschmann, Vladimír; Hon, Zdeněk

    2016-04-28

    Toxin weapon research, development, production and the ban on its uses is an integral part of international law, with particular attention paid to the protection against these weapons. In spite of this, hazards associated with toxins cannot be completely excluded. Some of these hazards are also pointed out in the present review. The article deals with the characteristics and properties of natural toxins and synthetic analogs potentially constituting the basis of toxin weapons. It briefly describes the history of military research and the use of toxins from distant history up to the present age. With respect to effective disarmament conventions, it mentions certain contemporary concepts of possible toxin applications for military purposes and the protection of public order (suppression of riots); it also briefly refers to the question of terrorism. In addition, it deals with certain traditional as well as modern technologies of the research, synthesis, and use of toxins, which can affect the continuing development of toxin weapons. These are, for example, cases of new toxins from natural sources, their chemical synthesis, production of synthetic analogs, the possibility of using methods of genetic engineering and modern biotechnologies or the possible applications of nanotechnology and certain pharmaceutical methods for the effective transfer of toxins into the organism. The authors evaluate the military importance of toxins based on their comparison with traditional chemical warfare agents. They appeal to the ethics of the scientific work as a principal condition for the prevention of toxin abuse in wars, military conflicts, as well as in non-military attacks.

  8. Botulinum toxin: bioweapon & magic drug.

    Science.gov (United States)

    Dhaked, Ram Kumar; Singh, Manglesh Kumar; Singh, Padma; Gupta, Pallavi

    2010-11-01

    Botulinum neurotoxins, causative agents of botulism in humans, are produced by Clostridium botulinum, an anaerobic spore-former Gram positive bacillus. Botulinum neurotoxin poses a major bioweapon threat because of its extreme potency and lethality; its ease of production, transport, and misuse; and the need for prolonged intensive care among affected persons. A single gram of crystalline toxin, evenly dispersed and inhaled, can kill more than one million people. The basis of the phenomenal potency of botulinum toxin is enzymatic; the toxin is a zinc proteinase that cleaves neuronal vesicle associated proteins responsible for acetylcholine release into the neuromuscular junction. As a military or terrorist weapon, botulinum toxin could be disseminated via aerosol or by contamination of water or food supplies, causing widespread casualties. A fascinating aspect of botulinum toxin research in recent years has been development of the most potent toxin into a molecule of significant therapeutic utility . It is the first biological toxin which is licensed for treatment of human diseases. In the late 1980s, Canada approved use of the toxin to treat strabismus, in 2001 in the removal of facial wrinkles and in 2002, the FDA in the United States followed suit. The present review focuses on both warfare potential and medical uses of botulinum neurotoxin.

  9. Different expression patterns of genes from the exo-xis region of bacteriophage λ and Shiga toxin-converting bacteriophage Ф24B following infection or prophage induction in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Sylwia Bloch

    Full Text Available Lambdoid bacteriophages serve as useful models in microbiological and molecular studies on basic biological process. Moreover, this family of viruses plays an important role in pathogenesis of enterohemorrhagic Escherichia coli (EHEC strains, as they are carriers of genes coding for Shiga toxins. Efficient expression of these genes requires lambdoid prophage induction and multiplication of the phage genome. Therefore, understanding the mechanisms regulating these processes appears essential for both basic knowledge and potential anti-EHEC applications. The exo-xis region, present in genomes of lambdoid bacteriophages, contains highly conserved genes of largely unknown functions. Recent report indicated that the Ea8.5 protein, encoded in this region, contains a newly discovered fused homeodomain/zinc-finger fold, suggesting its plausible regulatory role. Moreover, subsequent studies demonstrated that overexpression of the exo-xis region from a multicopy plasmid resulted in impaired lysogenization of E. coli and more effective induction of λ and Ф24B prophages. In this report, we demonstrate that after prophage induction, the increase in phage DNA content in the host cells is more efficient in E. coli bearing additional copies of the exo-xis region, while survival rate of such bacteria is lower, which corroborated previous observations. Importantly, by using quantitative real-time reverse transcription PCR, we have determined patterns of expressions of particular genes from this region. Unexpectedly, in both phages λ and Ф24B, these patterns were significantly different not only between conditions of the host cells infection by bacteriophages and prophage induction, but also between induction of prophages with various agents (mitomycin C and hydrogen peroxide. This may shed a new light on our understanding of regulation of lambdoid phage development, depending on the mode of lytic cycle initiation.

  10. Efficacy of a Blend of Sulfuric Acid and Sodium Sulfate against Shiga Toxin-Producing Escherichia coli, Salmonella, and Nonpathogenic Escherichia coli Biotype I on Inoculated Prerigor Beef Surface Tissue.

    Science.gov (United States)

    Scott-Bullard, Britteny R; Geornaras, Ifigenia; Delmore, Robert J; Woerner, Dale R; Reagan, James O; Morgan, J Bred; Belk, Keith E

    2017-12-01

    A study was conducted to investigate the efficacy of a sulfuric acid-sodium sulfate blend (SSS) against Escherichia coli O157:H7, non-O157 Shiga toxin-producing E. coli (STEC), Salmonella, and nonpathogenic E. coli biotype I on prerigor beef surface tissue. The suitability of using the nonpathogenic E. coli as a surrogate for in-plant validation studies was also determined by comparing the data obtained for the nonpathogenic inoculum with those for the pathogenic inocula. Prerigor beef tissue samples (10 by 10 cm) were inoculated (ca. 6 log CFU/cm 2 ) on the adipose side in a laboratory-scale spray cabinet with multistrain mixtures of E. coli O157:H7 (5 strains), non-O157 STEC (12 strains), Salmonella (6 strains), or E. coli biotype I (5 strains). Treatment parameters evaluated were two SSS pH values (1.5 and 1.0) and two spray application pressures (13 and 22 lb/in 2 ). Untreated inoculated beef tissue samples served as controls for initial bacterial populations. Overall, the SSS treatments lowered inoculated (6.1 to 6.4 log CFU/cm 2 ) bacterial populations by 0.6 to 1.5 log CFU/cm 2 (P SSS was applied to samples inoculated with any of the tested E. coli inocula; however, solution pH did have a significant effect (P SSS was applied to samples inoculated with Salmonella. Results indicated that the response of the nonpathogenic E. coli inoculum to the SSS treatments was similar (P ≥ 0.05) to that of the pathogenic inocula tested, making the E. coli biotype I strains viable surrogate organisms for in-plant validation of SSS efficacy on beef. The application of SSS at the tested parameters to prerigor beef surface tissue may be an effective intervention for controlling pathogens in a commercial beef harvest process.

  11. Enhanced sporulation and toxin production by a mutant derivative of ...

    African Journals Online (AJOL)

    fatima

    total proteins determined with the toxin producing organism. All values are the ... synthesis specific yield was the ratio of δ-endo-toxin (mg L-1) divided by .... corresponding to 31.8 mg. 108 spore. −1 delta-endotoxins, it became apparent that ...

  12. Toxin content and cytotoxicity of algal dietary supplements

    Energy Technology Data Exchange (ETDEWEB)

    Heussner, A.H.; Mazija, L. [Human and Environmental Toxicology, University of Konstanz, 78457 Konstanz (Germany); Fastner, J. [Federal Environmental Agency, Section II 3.3—Drinking-water resources and treatment, Berlin (Germany); Dietrich, D.R., E-mail: daniel.dietrich@uni-konstanz.de [Human and Environmental Toxicology, University of Konstanz, 78457 Konstanz (Germany)

    2012-12-01

    Blue-green algae (Spirulina sp., Aphanizomenon flos-aquae) and Chlorella sp. are commercially distributed as organic algae dietary supplements. Cyanobacterial dietary products in particular have raised serious concerns, as they appeared to be contaminated with toxins e.g. microcystins (MCs) and consumers repeatedly reported adverse health effects following consumption of these products. The aim of this study was to determine the toxin contamination and the in vitro cytotoxicity of algae dietary supplement products marketed in Germany. In thirteen products consisting of Aph. flos-aquae, Spirulina and Chlorella or mixtures thereof, MCs, nodularins, saxitoxins, anatoxin-a and cylindrospermopsin were analyzed. Five products tested in an earlier market study were re-analyzed for comparison. Product samples were extracted and analyzed for cytotoxicity in A549 cells as well as for toxin levels by (1) phosphatase inhibition assay (PPIA), (2) Adda-ELISA and (3) LC–MS/MS. In addition, all samples were analyzed by PCR for the presence of the mcyE gene, a part of the microcystin and nodularin synthetase gene cluster. Only Aph. flos-aquae products were tested positive for MCs as well as the presence of mcyE. The contamination levels of the MC-positive samples were ≤ 1 μg MC-LR equivalents g{sup −1} dw. None of the other toxins were found in any of the products. However, extracts from all products were cytotoxic. In light of the findings, the distribution and commercial sale of Aph. flos-aquae products, whether pure or mixed formulations, for human consumption appear highly questionable. -- Highlights: ► Marketed algae dietary supplements were analyzed for toxins. ► Methods: Phosphatase inhibition assay (PPIA), Adda-ELISA, LC-MS/MS. ► Aph. flos-aquae products all tested positive for microcystins. ► Products tested negative for nodularins, saxitoxins, anatoxin-a, cylindrospermopsin. ► Extracts from all products were cytotoxic.

  13. Influence of the vaginal microbiota on toxic shock syndrome toxin 1 production by Staphylococcus aureus.

    Science.gov (United States)

    MacPhee, Roderick A; Miller, Wayne L; Gloor, Gregory B; McCormick, John K; Hammond, Jo-Anne; Burton, Jeremy P; Reid, Gregor

    2013-03-01

    Menstrual toxic shock syndrome (TSS) is a serious illness that afflicts women of premenopausal age worldwide and arises from vaginal infection by Staphylococcus aureus and concurrent production of toxic shock syndrome toxin-1 (TSST-1). Studies have illustrated the capacity of lactobacilli to reduce S. aureus virulence, including the capacity to suppress TSST-1. We hypothesized that an aberrant microbiota characteristic of pathogenic bacteria would induce the increased production of TSST-1 and that this might represent a risk factor for the development of TSS. A S. aureus TSST-1 reporter strain was grown in the presence of vaginal swab contents collected from women with a clinically healthy vaginal status, women with an intermediate status, and those diagnosed with bacterial vaginosis (BV). Bacterial supernatant challenge assays were also performed to test the effects of aerobic vaginitis (AV)-associated pathogens toward TSST-1 production. While clinical samples from healthy and BV women suppressed toxin production, in vitro studies demonstrated that Streptococcus agalactiae and Enterococcus spp. significantly induced TSST-1 production, while some Lactobacillus spp. suppressed it. The findings suggest that women colonized by S. aureus and with AV, but not BV, may be more susceptible to menstrual TSS and would most benefit from prophylactic treatment.

  14. Plant Insecticidal Toxins in Ecological Networks

    Directory of Open Access Journals (Sweden)

    Sébastien Ibanez

    2012-04-01

    Full Text Available Plant secondary metabolites play a key role in plant-insect interactions, whether constitutive or induced, C- or N-based. Anti-herbivore defences against insects can act as repellents, deterrents, growth inhibitors or cause direct mortality. In turn, insects have evolved a variety of strategies to act against plant toxins, e.g., avoidance, excretion, sequestration and degradation of the toxin, eventually leading to a co-evolutionary arms race between insects and plants and to co-diversification. Anti-herbivore defences also negatively impact mutualistic partners, possibly leading to an ecological cost of toxin production. However, in other cases toxins can also be used by plants involved in mutualistic interactions to exclude inadequate partners and to modify the cost/benefit ratio of mutualism to their advantage. When considering the whole community, toxins have an effect at many trophic levels. Aposematic insects sequester toxins to defend themselves against predators. Depending on the ecological context, toxins can either increase insects’ vulnerability to parasitoids and entomopathogens or protect them, eventually leading to self-medication. We conclude that studying the community-level impacts of plant toxins can provide new insights into the synthesis between community and evolutionary ecology.

  15. Plant insecticidal toxins in ecological networks.

    Science.gov (United States)

    Ibanez, Sébastien; Gallet, Christiane; Després, Laurence

    2012-04-01

    Plant secondary metabolites play a key role in plant-insect interactions, whether constitutive or induced, C- or N-based. Anti-herbivore defences against insects can act as repellents, deterrents, growth inhibitors or cause direct mortality. In turn, insects have evolved a variety of strategies to act against plant toxins, e.g., avoidance, excretion, sequestration and degradation of the toxin, eventually leading to a co-evolutionary arms race between insects and plants and to co-diversification. Anti-herbivore defences also negatively impact mutualistic partners, possibly leading to an ecological cost of toxin production. However, in other cases toxins can also be used by plants involved in mutualistic interactions to exclude inadequate partners and to modify the cost/benefit ratio of mutualism to their advantage. When considering the whole community, toxins have an effect at many trophic levels. Aposematic insects sequester toxins to defend themselves against predators. Depending on the ecological context, toxins can either increase insects' vulnerability to parasitoids and entomopathogens or protect them, eventually leading to self-medication. We conclude that studying the community-level impacts of plant toxins can provide new insights into the synthesis between community and evolutionary ecology.

  16. Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins.

    Science.gov (United States)

    Mantzouki, Evanthia; Lürling, Miquel; Fastner, Jutta; de Senerpont Domis, Lisette; Wilk-Woźniak, Elżbieta; Koreivienė, Judita; Seelen, Laura; Teurlincx, Sven; Verstijnen, Yvon; Krztoń, Wojciech; Walusiak, Edward; Karosienė, Jūratė; Kasperovičienė, Jūratė; Savadova, Ksenija; Vitonytė, Irma; Cillero-Castro, Carmen; Budzyńska, Agnieszka; Goldyn, Ryszard; Kozak, Anna; Rosińska, Joanna; Szeląg-Wasielewska, Elżbieta; Domek, Piotr; Jakubowska-Krepska, Natalia; Kwasizur, Kinga; Messyasz, Beata; Pełechaty, Aleksandra; Pełechaty, Mariusz; Kokocinski, Mikolaj; García-Murcia, Ana; Real, Monserrat; Romans, Elvira; Noguero-Ribes, Jordi; Duque, David Parreño; Fernández-Morán, Elísabeth; Karakaya, Nusret; Häggqvist, Kerstin; Demir, Nilsun; Beklioğlu, Meryem; Filiz, Nur; Levi, Eti E.; Iskin, Uğur; Bezirci, Gizem; Tavşanoğlu, Ülkü Nihan; Özhan, Koray; Gkelis, Spyros; Panou, Manthos; Fakioglu, Özden; Avagianos, Christos; Kaloudis, Triantafyllos; Çelik, Kemal; Yilmaz, Mete; Marcé, Rafael; Catalán, Nuria; Bravo, Andrea G.; Buck, Moritz; Colom-Montero, William; Mustonen, Kristiina; Pierson, Don; Yang, Yang; Raposeiro, Pedro M.; Gonçalves, Vítor; Antoniou, Maria G.; Tsiarta, Nikoletta; McCarthy, Valerie; Perello, Victor C.; Feldmann, Tõnu; Laas, Alo; Panksep, Kristel; Tuvikene, Lea; Gagala, Ilona; Mankiewicz-Boczek, Joana; Yağcı, Meral Apaydın; Çınar, Şakir; Çapkın, Kadir; Yağcı, Abdulkadir; Cesur, Mehmet; Bilgin, Fuat; Bulut, Cafer; Uysal, Rahmi; Obertegger, Ulrike; Boscaini, Adriano; Flaim, Giovanna; Salmaso, Nico; Cerasino, Leonardo; Richardson, Jessica; Visser, Petra M.; Verspagen, Jolanda M. H.; Karan, Tünay; Soylu, Elif Neyran; Maraşlıoğlu, Faruk; Napiórkowska-Krzebietke, Agnieszka; Ochocka, Agnieszka; Pasztaleniec, Agnieszka; Antão-Geraldes, Ana M.; Vasconcelos, Vitor; Morais, João; Vale, Micaela; Köker, Latife; Akçaalan, Reyhan; Albay, Meriç; Špoljarić Maronić, Dubravka; Stević, Filip; Žuna Pfeiffer, Tanja; Fonvielle, Jeremy; Straile, Dietmar; Rothhaupt, Karl-Otto; Hansson, Lars-Anders; Urrutia-Cordero, Pablo; Bláha, Luděk; Geriš, Rodan; Fránková, Markéta; Koçer, Mehmet Ali Turan; Alp, Mehmet Tahir; Remec-Rekar, Spela; Elersek, Tina; Triantis, Theodoros; Zervou, Sevasti-Kiriaki; Hiskia, Anastasia; Haande, Sigrid; Skjelbred, Birger; Madrecka, Beata; Nemova, Hana; Drastichova, Iveta; Chomova, Lucia; Edwards, Christine; Sevindik, Tuğba Ongun; Tunca, Hatice; Önem, Burçin; Aleksovski, Boris; Krstić, Svetislav; Vucelić, Itana Bokan; Nawrocka, Lidia; Salmi, Pauliina; Machado-Vieira, Danielle; de Oliveira, Alinne Gurjão; Delgado-Martín, Jordi; García, David; Cereijo, Jose Luís; Gomà, Joan; Trapote, Mari Carmen; Vegas-Vilarrúbia, Teresa; Obrador, Biel; Grabowska, Magdalena; Karpowicz, Maciej; Chmura, Damian; Úbeda, Bárbara; Gálvez, José Ángel; Özen, Arda; Christoffersen, Kirsten Seestern; Warming, Trine Perlt; Kobos, Justyna; Mazur-Marzec, Hanna; Pérez-Martínez, Carmen; Ramos-Rodríguez, Eloísa; Arvola, Lauri; Alcaraz-Párraga, Pablo; Toporowska, Magdalena; Pawlik-Skowronska, Barbara; Niedźwiecki, Michał; Pęczuła, Wojciech; Leira, Manel; Hernández, Armand; Moreno-Ostos, Enrique; Blanco, José María; Rodríguez, Valeriano; Montes-Pérez, Jorge Juan; Palomino, Roberto L.; Rodríguez-Pérez, Estela; Carballeira, Rafael; Camacho, Antonio; Picazo, Antonio; Rochera, Carlos; Santamans, Anna C.; Ferriol, Carmen; Romo, Susana; Soria, Juan Miguel; Dunalska, Julita; Sieńska, Justyna; Szymański, Daniel; Kruk, Marek; Kostrzewska-Szlakowska, Iwona; Jasser, Iwona; Žutinić, Petar; Gligora Udovič, Marija; Plenković-Moraj, Anđelka; Frąk, Magdalena; Bańkowska-Sobczak, Agnieszka; Wasilewicz, Michał; Özkan, Korhan; Maliaka, Valentini; Kangro, Kersti; Grossart, Hans-Peter; Paerl, Hans W.; Carey, Cayelan C.; Ibelings, Bas W.

    2018-04-13

    Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins). Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a) and cytotoxins (e.g., cylindrospermopsin) due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and light. In summer 2015, we collected samples across Europe to investigate the effect of nutrient and temperature gradients on the variability of toxin production at a continental scale. Direct and indirect effects of temperature were the main drivers of the spatial distribution in the toxins produced by the cyanobacterial community, the toxin concentrations and toxin quota. Generalized linear models showed that a Toxin Diversity Index (TDI) increased with latitude, while it decreased with water stability. Increases in TDI were explained through a significant increase in toxin variants such as MC-YR, anatoxin and cylindrospermopsin, accompanied by a decreasing presence of MC-LR. While global warming continues, the direct and indirect effects of increased lake temperatures will drive changes in the distribution of cyanobacterial toxins in Europe, potentially promoting selection of a few highly toxic species or strains.

  17. Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins

    Directory of Open Access Journals (Sweden)

    Evanthia Mantzouki

    2018-04-01

    Full Text Available Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins. Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a and cytotoxins (e.g., cylindrospermopsin due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and light. In summer 2015, we collected samples across Europe to investigate the effect of nutrient and temperature gradients on the variability of toxin production at a continental scale. Direct and indirect effects of temperature were the main drivers of the spatial distribution in the toxins produced by the cyanobacterial community, the toxin concentrations and toxin quota. Generalized linear models showed that a Toxin Diversity Index (TDI increased with latitude, while it decreased with water stability. Increases in TDI were explained through a significant increase in toxin variants such as MC-YR, anatoxin and cylindrospermopsin, accompanied by a decreasing presence of MC-LR. While global warming continues, the direct and indirect effects of increased lake temperatures will drive changes in the distribution of cyanobacterial toxins in Europe, potentially promoting selection of a few highly toxic species or strains.

  18. Dinophysis Toxins: Causative Organisms, Distribution and Fate in Shellfish

    Science.gov (United States)

    Reguera, Beatriz; Riobó, Pilar; Rodríguez, Francisco; Díaz, Patricio A.; Pizarro, Gemita; Paz, Beatriz; Franco, José M.; Blanco, Juan

    2014-01-01

    Several Dinophysis species produce diarrhoetic toxins (okadaic acid and dinophysistoxins) and pectenotoxins, and cause gastointestinal illness, Diarrhetic Shellfish Poisoning (DSP), even at low cell densities (Chile, and Europe. Toxicity and toxin profiles are very variable, more between strains than species. The distribution of DSP events mirrors that of shellfish production areas that have implemented toxin regulations, otherwise misinterpreted as bacterial or viral contamination. Field observations and laboratory experiments have shown that most of the toxins produced by Dinophysis are released into the medium, raising questions about the ecological role of extracelular toxins and their potential uptake by shellfish. Shellfish contamination results from a complex balance between food selection, adsorption, species-specific enzymatic transformations, and allometric processes. Highest risk areas are those combining Dinophysis strains with high cell content of okadaates, aquaculture with predominance of mytilids (good accumulators of toxins), and consumers who frequently include mussels in their diet. Regions including pectenotoxins in their regulated phycotoxins will suffer from much longer harvesting bans and from disloyal competition with production areas where these toxins have been deregulated. PMID:24447996

  19. ChLae1 and ChVel1 regulate T-toxin production, virulence, oxidative stress response, and development of the maize pathogen Cochliobolus heterostrophus.

    Directory of Open Access Journals (Sweden)

    Dongliang Wu

    2012-02-01

    Full Text Available LaeA and VeA coordinate secondary metabolism and differentiation in response to light signals in Aspergillus spp. Their orthologs, ChLae1 and ChVel1, were identified in the maize pathogen Cochliobolus heterostrophus, known to produce a wealth of secondary metabolites, including the host selective toxin, T-toxin. Produced by race T, T-toxin promotes high virulence to maize carrying Texas male sterile cytoplasm (T-cms. T-toxin production is significantly increased in the dark in wild type (WT, whereas Chvel1 and Chlae1 mutant toxin levels are much reduced in the dark compared to WT. Correspondingly, expression of T-toxin biosynthetic genes (Tox1 is up-regulated in the dark in WT, while dark-induced expression is much reduced/minimal in Chvel1 and Chlae1 mutants. Toxin production and Tox1 gene expression are increased in ChVEL1 overexpression (OE strains grown in the dark and in ChLAE1 strains grown in either light or dark, compared to WT. These observations establish ChLae1 and ChVel1 as the first factors known to regulate host selective toxin production. Virulence of Chlae1 and Chvel1 mutants and OE strains is altered on both T-cms and normal cytoplasm maize, indicating that both T-toxin mediated super virulence and basic pathogenic ability are affected. Deletion of ChLAE1 or ChVEL1 reduces tolerance to H(2O(2. Expression of CAT3, one of the three catalase genes, is reduced in the Chvel1 mutant. Chlae1 and Chvel1 mutants also show decreased aerial hyphal growth, increased asexual sporulation and female sterility. ChLAE1 OE strains are female sterile, while ChVEL1 OE strains are more fertile than WT. ChLae1 and ChVel1 repress expression of 1,8-dihydroxynaphthalene (DHN melanin biosynthesis genes, and, accordingly, melanization is enhanced in Chlae1 and Chvel1 mutants, and reduced in OE strains. Thus, ChLae1 and ChVel1 positively regulate T-toxin biosynthesis, pathogenicity and super virulence, oxidative stress responses, sexual development, and

  20. Production of T-2 toxin and deoxynivalenol in the presence of different disinfectants

    OpenAIRE

    Dana Hrubošová; Jarmila Vytřasová; Iveta Brožková

    2015-01-01

    The aim of the work was to examine the effect of different disinfectants on production trichothecenes (especially of T-2 toxin and deoxynivalenol). Lipophilicity, chemical structure, the presence of bioactive groups and functional groups in their structure modifies biological activity and toxic potency of trichothecenes. For this reason, limits have been established designating maximum levels of mycotoxins in cereals while maintaining proper growing practices. Appropriate nutr...

  1. Effect of Sodium Nitrite and Sodium Nitrate on Botulinal Toxin Production and Nitrosamine Formation in Wieners

    Science.gov (United States)

    Hustad, Gerald O.; Cerveny, John G.; Trenk, Hugh; Deibel, Robert H.; Kautter, Donald A.; Fazio, Thomas; Johnston, Ralph W.; Kolari, Olaf E.

    1973-01-01

    Wieners were formulated and processed approximating commercial conditions as closely as possible. Twenty-four batches of product were made with the addition of six levels of sodium nitrite (0, 50, 100, 150, 200, and 300 μg/g), four levels of sodium nitrate (0, 50, 150, and 450 μg/g), and two levels of Clostridium botulinum (0 and 620 spores/g). After formulation, processing, and vacuum packaging, portions of each batch were incubated at 27 C or held for 21 days at 7 C followed by incubation at 27 C for 56 days. The latter storage condition approximated distribution of product through commercial channels and potential temperature abuse at the consumer level. Samples were analyzed for botulinal toxin, nitrite, and nitrate levels after 3, 7, 14, 21, 28, and 56 days of incubation. When nitrite was not added, toxic samples were detected after 14 days of incubation at 27 C. At the lowest level of nitrite added (50 μg/g), no toxic samples were observed until 56 days of incubation. Higher levels of nitrite completely inhibited toxin production throughout the incubation period. Nine uninoculated samples, representing various levels and combinations of nitrite and nitrate, were evaluated organoleptically. The flavor quality of wieners made with nitrite was judged significantly higher (P = 0.05) than of wieners made without nitrite. The nine samples were negative for 14 volatile nitrosamines at a sensitivity level of 10 ng/g. The results indicated that nitrite effectively inhibited botulinal toxin formation at commercially employed levels in wieners and that detectable quantities of nitrosamines were not produced during preparation and processing of the product for consumption. PMID:4580194

  2. Comparing two botulinum toxin type A formulations using manufacturers' product summaries.

    Science.gov (United States)

    Wenzel, R; Jones, D; Borrego, J A

    2007-08-01

    Because of the unique pharmacology and clinical versatility of botulinum toxin (BoNT), particularly BoNT serotype A (BoNTA), a need exists for discussion of the current data on similarities and differences between two BoNTA products, BOTOX and Dysport. We compared the physiochemical and pharmacological properties of BOTOX and Dysport using information from the Summary of Product Characteristics (SmPC) documents from a number of countries around the world. Our analysis based on the SmPC documents demonstrated distinct differences in physical characteristics, breadth of approved indications, dosing and administration, and the incidence and severity of adverse events. BOTOX and Dysport are not bioequivalent. Many of the differences between BOTOX and Dysport discussed within are probably related to the differences in their physical characteristics.

  3. Isolation and characterization of non-O157 Shiga toxin-producing Escherichia coli from beef carcasses, cuts and trimmings of abattoirs in Argentina

    Science.gov (United States)

    Brusa, Victoria; Restovich, Viviana; Galli, Lucía; Teitelbaum, David; Signorini, Marcelo; Brasesco, Hebe; Londero, Alejandra; García, Diego; Padola, Nora Lía; Superno, Valeria; Sanz, Marcelo; Petroli, Sandra; Costa, Magdalena; Bruzzone, Mariana; Sucari, Adriana; Ferreghini, Marcela; Linares, Luciano; Suberbie, Germán; Rodríguez, Ricardo

    2017-01-01

    Several foods contaminated with Shiga toxin-producing Escherichia coli (STEC) are associated with human diseases. Some countries have established microbiological criteria for non-O157 STEC, thus, the absence of serogroups O26, O45, O103, O104, O111, O121, and O145 in sprouts from the European Union or ground beef and beef trimmings from the United States is mandatory. While in Argentina screening for O26, O103, O111, O145 and O121 in ground beef, ready-to-eat food, sausages and vegetables is mandatory, other countries have zero-tolerance for all STEC in chilled beef. The aim of this study was to provide data on the prevalence of non-O157 STEC isolated from beef processed in eight Argentinean cattle slaughterhouses producing beef for export and local markets, and to know the non-O157 STEC profiles through strain characterization and genotypic analysis. Samples (n = 15,965) from 3,205 beef carcasses, 9,570 cuts and 3,190 trimmings collected between March and September 2014 were processed in pools of five samples each. Pools of samples (n = 3,193) from 641 carcasses, 1,914 cuts and 638 trimming were analyzed for non-O157 STEC isolation according to ISO/CEN 13136:2012. Of these, 37 pools of carcasses (5.8%), 111 pools of cuts (5.8%) and 45 pools of trimmings (7.0%) were positive for non-O157 STEC. STEC strains (n = 200) were isolated from 193 pools of samples. The most prevalent serotypes were O174:H21, O185:H7, O8:H19, O178:H19 and O130:H11, and the most prevalent genotypes were stx2c(vh-b) and stx2a/saa/ehxA. O103:H21 strain was eae-positive and one O178:H19 strain was aggR/aaiC-positive. The prevalence of non-O157 STEC in beef carcasses reported here was low. None of the non-O157 STEC strains isolated corresponded to the non-O157 STEC serotypes and virulence profiles isolated from human cases in Argentina in the same study period. The application of microbiological criteria for each foodstuff should be determined by risk analysis in order to have a stringent

  4. Application of whole genome sequence data in analyzing the molecular epidemiology of Shiga toxin-producing Escherichia coli O157:H7/H.

    Science.gov (United States)

    Yokoyama, Eiji; Hirai, Shinichiro; Ishige, Taichiro; Murakami, Satoshi

    2018-01-02

    Seventeen clusters of Shiga toxin-producing Escherichia coli O157:H7/- (O157) strains, determined by cluster analysis of pulsed-field gel electrophoresis patterns, were analyzed using whole genome sequence (WGS) data to investigate this pathogen's molecular epidemiology. The 17 clusters included 136 strains containing strains from nine outbreaks, with each outbreak caused by a single source contaminated with the organism, as shown by epidemiological contact surveys. WGS data of these strains were used to identify single nucleotide polymorphisms (SNPs) by two methods: short read data were directly mapped to a reference genome (mapping derived SNPs) and common SNPs between the mapping derived SNPs and SNPs in assembled data of short read data (common SNPs). Among both SNPs, those that were detected in genes with a gap were excluded to remove ambiguous SNPs from further analysis. The effectiveness of both SNPs was investigated among all the concatenated SNPs that were detected (whole SNP set); SNPs were divided into three categories based on the genes in which they were located (i.e., backbone SNP set, O-island SNP set, and mobile element SNP set); and SNPs in non-coding regions (intergenic region SNP set). When SNPs from strains isolated from the nine single source derived outbreaks were analyzed using an unweighted pair group method with arithmetic mean tree (UPGMA) and a minimum spanning tree (MST), the maximum pair-wise distances of the backbone SNP set of the mapping derived SNPs were significantly smaller than those of the whole and intergenic region SNP set on both UPGMAs and MSTs. This significant difference was also observed when the backbone SNP set of the common SNPs were examined (Steel-Dwass test, P≤0.01). When the maximum pair-wise distances were compared between the mapping derived and common SNPs, significant differences were observed in those of the whole, mobile element, and intergenic region SNP set (Wilcoxon signed rank test, P≤0.01). When all

  5. Isolation and characterization of non-O157 Shiga toxin-producing Escherichia coli from beef carcasses, cuts and trimmings of abattoirs in Argentina.

    Directory of Open Access Journals (Sweden)

    Victoria Brusa

    Full Text Available Several foods contaminated with Shiga toxin-producing Escherichia coli (STEC are associated with human diseases. Some countries have established microbiological criteria for non-O157 STEC, thus, the absence of serogroups O26, O45, O103, O104, O111, O121, and O145 in sprouts from the European Union or ground beef and beef trimmings from the United States is mandatory. While in Argentina screening for O26, O103, O111, O145 and O121 in ground beef, ready-to-eat food, sausages and vegetables is mandatory, other countries have zero-tolerance for all STEC in chilled beef. The aim of this study was to provide data on the prevalence of non-O157 STEC isolated from beef processed in eight Argentinean cattle slaughterhouses producing beef for export and local markets, and to know the non-O157 STEC profiles through strain characterization and genotypic analysis. Samples (n = 15,965 from 3,205 beef carcasses, 9,570 cuts and 3,190 trimmings collected between March and September 2014 were processed in pools of five samples each. Pools of samples (n = 3,193 from 641 carcasses, 1,914 cuts and 638 trimming were analyzed for non-O157 STEC isolation according to ISO/CEN 13136:2012. Of these, 37 pools of carcasses (5.8%, 111 pools of cuts (5.8% and 45 pools of trimmings (7.0% were positive for non-O157 STEC. STEC strains (n = 200 were isolated from 193 pools of samples. The most prevalent serotypes were O174:H21, O185:H7, O8:H19, O178:H19 and O130:H11, and the most prevalent genotypes were stx2c(vh-b and stx2a/saa/ehxA. O103:H21 strain was eae-positive and one O178:H19 strain was aggR/aaiC-positive. The prevalence of non-O157 STEC in beef carcasses reported here was low. None of the non-O157 STEC strains isolated corresponded to the non-O157 STEC serotypes and virulence profiles isolated from human cases in Argentina in the same study period. The application of microbiological criteria for each foodstuff should be determined by risk analysis in order to have a

  6. Influence of the Vaginal Microbiota on Toxic Shock Syndrome Toxin 1 Production by Staphylococcus aureus

    OpenAIRE

    MacPhee, Roderick A.; Miller, Wayne L.; Gloor, Gregory B.; McCormick, John K.; Hammond, Jo-Anne; Burton, Jeremy P.; Reid, Gregor

    2013-01-01

    Menstrual toxic shock syndrome (TSS) is a serious illness that afflicts women of premenopausal age worldwide and arises from vaginal infection by Staphylococcus aureus and concurrent production of toxic shock syndrome toxin-1 (TSST-1). Studies have illustrated the capacity of lactobacilli to reduce S. aureus virulence, including the capacity to suppress TSST-1. We hypothesized that an aberrant microbiota characteristic of pathogenic bacteria would induce the increased production of TSST-1 and...

  7. Prevalence of Stx-producing Shigella species isolated from French Travelers Returning from the Caribbean: An Emerging Pathogen with International Implications

    Science.gov (United States)

    Gray, Miranda D.; Lacher, David W.; Leonard, Susan R.; Abbott, Jason; Zhao, Shaohua; Lampel, Keith A.; Prothery, Estelle; Gouali, Malika; Weill, François-Xavier; Maurelli, Anthony T.

    2015-01-01

    Shiga toxins are potent cytotoxins that inhibit host cell protein synthesis, leading to cell death. Classically, these toxins are associated with intestinal infections due to Shiga toxin-producing Escherichia coli or Shigella dysenteriae serotype 1 and infections with these strains can lead to hemolytic uremic syndrome. Over the past decade there is increasing recognition that Shiga toxin is produced by additional Shigella species. We recently reported the presence and expression of stx genes in Shigella flexneri 2a clinical isolates. The toxin genes were carried by a new stx-encoding bacteriophage and infection with these strains correlated with recent travel to Haiti or the Dominican Republic. In this study we further explored the epidemiological link to this region by utilizing the French National Reference Center for Escherichia coli, Shigella and Salmonella collection to survey the frequency of Stx-producing Shigella species isolated from French travelers returning from the Caribbean. About 21% of the isolates tested were found to encode and produce Stx. These isolates included strains of S. flexneri 2a, S. flexneri Y, and S. dysenteriae 4. All of the travelers whom were infected with Stx-producing Shigella had recently traveled to Haiti, the Dominican Republic, or French Guiana. Furthermore, whole genome sequencing found that the toxin genes were encoded by a prophage that was highly identical to the phage we identified in our previous study. These findings demonstrate that this new stx-encoding prophage is circulating within that geographical area, has spread to other continents, and is capable of spreading to multiple Shigella serogroups. PMID:25980352

  8. Aislamiento de Escherichia coli productor de toxina Shiga durante un brote de gastroenteritis en un Jardín Maternal de la Ciudad de Mar del Plata Isolation of Shiga toxin-producing Escherichia coli strains during a gastrointestinal outbreak at a day care center in Mar del Plata City

    Directory of Open Access Journals (Sweden)

    D. Gomez

    2005-12-01

    considered risk factors. One case had Shiga toxin-producing Escherichia coli (STEC O103:H2 infection and other STEC O26:H11.The duration of shedding for the child with O26:H11 infection was 37 days. In the other symptomatic children, the pathogen was not recovered from fecal samples collected 6 or more days after the onset of the illness. This emphasizes that the collection of early samples is necessary to recover STEC strains. In order to prevent and control enteric diseases in day care facilities the following measures are necessary: optimal hygiene standards, early case reporting, and exclusion of those who remain culture-positive.

  9. A 3D intestinal tissue model supports Clostridioides difficile germination, colonization, toxin production and epithelial damage.

    Science.gov (United States)

    Shaban, Lamyaa; Chen, Ying; Fasciano, Alyssa C; Lin, Yinan; Kaplan, David L; Kumamoto, Carol A; Mecsas, Joan

    2018-04-01

    Endospore-forming Clostridioides difficile is a causative agent of antibiotic-induced diarrhea, a major nosocomial infection. Studies of its interactions with mammalian tissues have been hampered by the fact that C. difficile requires anaerobic conditions to survive after spore germination. We recently developed a bioengineered 3D human intestinal tissue model and found that low O 2 conditions are produced in the lumen of these tissues. Here, we compared the ability of C. difficile spores to germinate, produce toxin and cause tissue damage in our bioengineered 3D tissue model versus in a 2D transwell model in which human cells form a polarized monolayer. 3D tissue models or 2D polarized monolayers on transwell filters were challenged with the non-toxin producing C. difficile CCUG 37787 serotype X (ATCC 43603) and the toxin producing UK1 C. difficile spores in the presence of the germinant, taurocholate. Spores germinated in both the 3D tissue model as well as the 2D transwell system, however toxin activity was significantly higher in the 3D tissue models compared to the 2D transwells. Moreover, the epithelium damage in the 3D tissue model was significantly more severe than in 2D transwells and damage correlated significantly with the level of toxin activity detected but not with the amount of germinated spores. Combined, these results show that the bioengineered 3D tissue model provides a powerful system with which to study early events leading to toxin production and tissue damage of C. difficile with mammalian cells under anaerobic conditions. Furthermore, these systems may be useful for examining the effects of microbiota, novel drugs and other potential therapeutics directed towards C. difficile infections. Copyright © 2018 Elsevier Ltd. All rights reserved.

  10. Effect of severe weather events on the shedding of Shiga toxigenic Escherichia coli in slaughter cattle and phenotype of serogroup O157 isolates.

    Science.gov (United States)

    Stanford, Kim; Reuter, Tim; Bach, Susan J; Chui, Linda; Ma, Angela; Conrad, Cheyenne C; Tostes, Renata; McAllister, Tim A

    2017-09-01

    High-event periods (HEPs) occur sporadically when beef carcasses and meat have episodes of acute contamination with Shiga toxin-producing Escherichia coli (STEC). In this study, severe weather events were investigated as catalysts for HEPs based on PCR and isolate prevalence of seven E. coli serogroups in slaughter cattle feces. Winter ambient temperatures with daily means 10.5oC warmer or 12.3°C colder than seasonal norms (-10.4°C) most altered STEC shedding. Fecal samples yielded increased proportions (P  10 min and one also had strong biofilm-forming potential. However, this isolate lacked eae and stx genes. Severe weather can influence STEC shedding, particularly of O157, and could possibly trigger HEPs. However, our data suggest that it is unlikely for isolates to carry virulence genes and possess phenotypes capable of evading post-harvest microbiological interventions. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Cellular trafficking of quantum dot-ligand bioconjugates and their induction of changes in normal routing of unconjugated ligands

    DEFF Research Database (Denmark)

    Tekle, Christina; van Deurs, Bo; Sandvig, Kirsten

    2008-01-01

    Can quantum dots (Qdots) act as relevant intracellular probes to investigate routing of ligands in live cells? The intracellular trafficking of Qdots that were coupled to the plant toxin ricin, Shiga toxin, or the ligand transferrin (Tf) was studied by confocal fluorescence microscopy. The Tf...

  12. ArgO145, a Stx2a prophage of a bovine O145 : H-STEC strain, is closely related to phages of virulent human strains

    NARCIS (Netherlands)

    Krüger, A; Burgán, J; Friedrich, A W; Jwa, Rossen; Pma, Lucchesi

    Shiga toxins (Stx) are the main virulence factor of a pathogroup of Escherichia coli strains that cause severe human diseases. These toxins are encoded in prophages (Stx prophages), and generally their expression depends on prophage induction. Several studies have reported high diversity among both

  13. Associations Between Hydration Status, Intravenous Fluid Administration, and Outcomes of Patients Infected With Shiga Toxin-Producing Escherichia coli: A Systematic Review and Meta-analysis.

    Science.gov (United States)

    Grisaru, Silviu; Xie, Jianling; Samuel, Susan; Hartling, Lisa; Tarr, Phillip I; Schnadower, David; Freedman, Stephen B

    2017-01-01

    The associations between hydration status, intravenous fluid administration, and outcomes of patients infected with Shiga toxin-producing Escherichia coli (STEC) remain unclear. To determine the relationship between hydration status, the development and severity of hemolytic uremic syndrome (HUS), and adverse outcomes in STEC-infected individuals. MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials via the OvidSP platform, PubMed via the National Library of Medicine, CINAHL Plus with full text, Scopus, Web of Science, ClinicalTrials.gov, reference lists, and gray literature were systematically searched. Two reviewers independently identified studies that included patients with hydration status documentation, proven or presumed STEC infection, and some form of HUS that developed. No language restrictions were applied. Two reviewers independently extracted individual study data, including study characteristics, population, and outcomes. Risk of bias was assessed using the Newcastle-Ottawa Scale; strength of evidence was adjudicated using the Grading of Recommendations Assessment, Development, and Evaluation method. Meta-analyses were conducted using random-effects models. Development of HUS, complications (ie, oligoanuric renal failure, involvement of the central nervous system, or death), and interventions (ie, renal replacement therapy). Eight studies comprising 1511 patients (all children) met eligibility criteria. Unpublished data were provided by the authors of 7 published reports. The median risk-of-bias score was 7.5 (range, 6-9). No studies evaluated the effect of hydration during STEC infections on the risk for HUS. A hematocrit value greater than 23% as a measure of hydration status at presentation with HUS was associated with the development of oligoanuric HUS (OR, 2.38 [95% CI, 1.30-4.35]; I2 = 2%), renal replacement therapy (OR, 1.90 [95% CI, 1.25-2.90]; I2 = 17%), and death (OR, 5.13 [95% CI, 1.50-17.57]; I2 = 55%). Compared with

  14. Longitudinal study of two Irish dairy herds: Low numbers of of Shiga toxin-producing Escherichia coli O157 and O26 super-shedders identified

    Directory of Open Access Journals (Sweden)

    Brenda Patricia Murphy

    2016-11-01

    Full Text Available A 12-month longitudinal study was undertaken on two dairy herds to ascertain the Shiga-toxin producing Escherichia coli (STEC O157 and O26 shedding status of the animals and its impact (if any on raw milk. Cattle are a recognised reservoir for these organisms with associated public health and environmental implications. Animals shedding E. coli O157 at >10,000 CFU/g of faeces have been deemed super-shedders. There is a gap in the knowledge regarding super-shedding of other STEC serogroups. A cohort of 40 lactating cows from herds previously identified as positive for VTEC in a national surveillance project were sampled every second month between August, 2013 and July, 2014. Metadata on any potential super-shedders was documented including e.g. age of the animal, number of lactations and days in lactation, nutritional condition, somatic cell count and content of protein in milk to assess if any were associated with risk factors for super-shedding. Recto-anal mucosal swabs, raw milk, milk filters and water samples were procured for each herd. The swabs were examined for E. coli O157 and O26 using a quantitative real time PCR method. Counts (CFU swab-1 were obtained from a standard calibration curve that related real-time PCR cycle threshold (Ct values against the initial concentration of O157 or O26 in the samples. Results from Farm A: 305 animals were analysed; 15 E. coli O157 (5% were recovered, 13 were denoted STEC encoding either stx1 and/or stx2 virulence genes and 5 (2% STEC O26 were recovered. One super-shedder was identified shedding STEC O26 (stx1&2. Farm B: 224 animals were analysed; eight E. coli O157 (3.5% were recovered (seven were STEC and 9 (4% STEC O26 were recovered. Three super-shedders were identified, one was shedding STEC O157 (stx2 and two STEC O26 (stx2. Three encoded the adhering and effacement gene (eae and one isolate additionally encoded the haemolysin gene (hlyA. The results of this study show, low numbers of super

  15. The Gene tia, Harbored by the Subtilase-Encoding Pathogenicity Island, Is Involved in the Ability of Locus of Enterocyte Effacement-Negative Shiga Toxin-Producing Escherichia coli Strains To Invade Monolayers of Epithelial Cells

    Science.gov (United States)

    Chiani, Paola; Michelacci, Valeria; Minelli, Fabio; Caprioli, Alfredo; Morabito, Stefano

    2017-01-01

    ABSTRACT Locus of enterocyte effacement (LEE)-negative Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are human pathogens that lack the LEE locus, a pathogenicity island (PAI) involved in the intimate adhesion of LEE-positive strains to the host gut epithelium. The mechanism used by LEE-negative STEC strains to colonize the host intestinal mucosa is still not clear. The cell invasion determinant tia, previously described in enterotoxigenic E. coli strains, has been identified in LEE-negative STEC strains that possess the subtilase-encoding pathogenicity island (SE-PAI). We evaluated the role of the gene tia, present in these LEE-negative STEC strains, in the invasion of monolayers of cultured cells. We observed that these strains were able to invade Caco-2 and HEp-2 cell monolayers and compared their invasion ability with that of a mutant strain in which the gene tia had been inactivated. Mutation of the gene tia resulted in a strong reduction of the invasive phenotype, and complementation of the tia mutation with a functional copy of the gene restored the invasion activity. Moreover, we show that the gene tia is overexpressed in bacteria actively invading cell monolayers, demonstrating that tia is involved in the ability to invade cultured monolayers of epithelial cells shown by SE-PAI-positive E. coli, including STEC, strains. However, the expression of the tia gene in the E. coli K-12 strain JM109 was not sufficient, in its own right, to confer to this strain the ability to invade cell monolayers, suggesting that at least another factor must be involved in the invasion ability displayed by the SE-PAI-positive strains. PMID:28893912

  16. The Gene tia, Harbored by the Subtilase-Encoding Pathogenicity Island, Is Involved in the Ability of Locus of Enterocyte Effacement-Negative Shiga Toxin-Producing Escherichia coli Strains To Invade Monolayers of Epithelial Cells.

    Science.gov (United States)

    Bondì, Roslen; Chiani, Paola; Michelacci, Valeria; Minelli, Fabio; Caprioli, Alfredo; Morabito, Stefano

    2017-12-01

    Locus of enterocyte effacement (LEE)-negative Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are human pathogens that lack the LEE locus, a pathogenicity island (PAI) involved in the intimate adhesion of LEE-positive strains to the host gut epithelium. The mechanism used by LEE-negative STEC strains to colonize the host intestinal mucosa is still not clear. The cell invasion determinant tia , previously described in enterotoxigenic E. coli strains, has been identified in LEE-negative STEC strains that possess the subtilase-encoding pathogenicity island (SE-PAI). We evaluated the role of the gene tia , present in these LEE-negative STEC strains, in the invasion of monolayers of cultured cells. We observed that these strains were able to invade Caco-2 and HEp-2 cell monolayers and compared their invasion ability with that of a mutant strain in which the gene tia had been inactivated. Mutation of the gene tia resulted in a strong reduction of the invasive phenotype, and complementation of the tia mutation with a functional copy of the gene restored the invasion activity. Moreover, we show that the gene tia is overexpressed in bacteria actively invading cell monolayers, demonstrating that tia is involved in the ability to invade cultured monolayers of epithelial cells shown by SE-PAI-positive E. coli , including STEC, strains. However, the expression of the tia gene in the E. coli K-12 strain JM109 was not sufficient, in its own right, to confer to this strain the ability to invade cell monolayers, suggesting that at least another factor must be involved in the invasion ability displayed by the SE-PAI-positive strains. Copyright © 2017 American Society for Microbiology.

  17. Microbiological analysis of pre-packed sweet basil (Ocimum basilicum) and coriander (Coriandrum sativum) leaves for the presence of Salmonella spp. and Shiga toxin-producing E. coli.

    Science.gov (United States)

    Delbeke, Stefanie; Ceuppens, Siele; Jacxsens, Liesbeth; Uyttendaele, Mieke

    2015-09-02

    Enteric pathogens, such as Salmonella spp. and pathogenic Escherichia coli, have been detected and associated with food borne outbreaks from (imported) fresh leafy herbs. Screening on imported herbs from South East Asian countries has been described. However, limited information on prevalence of these pathogens is available from other sourcing regions. Therefore, fresh pre-packed basil and coriander leaves from a Belgian trading company were investigated for the presence of Salmonella spp., Shiga toxin-producing E. coli (STEC), generic E. coli and coliforms. In total 592 samples were collected originating from Belgium, Israel and Cyprus during 2013-2014. Multiplex PCR followed by further culture confirmation was used for the detection of Salmonella spp. and STEC, whereas the Petrifilm Select E. coli and VRBL-agar were used, respectively, for the enumeration of E. coli and coliforms. Salmonella was detected in 10 out of 592 samples (25g) (1.7%; 5 from basil and 5 from coriander), of which two samples were sourced from Israel and eight from Cyprus. The presence of STEC was suspected in 11 out of 592 samples (25g) (1.9%; 3 basil and 8 coriander), due to the detection of stx and eae genes, of which one sample originated from Belgium, four from Israel and six from Cyprus. No STEC was isolated by culture techniques, but in three samples a serotype (O26, O103 or O111) with its most likely associated eae-variant (β or θ) was detected by PCR. Generic E. coli was enumerated in 108 out of 592 samples, whereby 55, 32 and 13 samples respectively between 10-100, 100-1000 and 1000-10,000cfu/g and 8 samples exceeding 10,000cfu/g. Coliforms were enumerated in all herb samples at variable levels ranging from 1.6 to 7.5logcfu/g. Further statistics indicate that the E. coli class (categorized by level) was significantly correlated with the presence of Salmonella (pbasil and coriander sourced from different cultivation regions, may contain enteric pathogens and potentially pose a

  18. Effect of low-dose irradiation on growth of and toxin production by Staphylococcus aureus and Bacillus cereus in roast beef and gravy.

    Science.gov (United States)

    Grant, I R; Nixon, C R; Patterson, M F

    1993-03-01

    The effect of irradiation (2 kGy) on growth of and toxin production by Staphylococcus aureus and Bacillus cereus in roast beef and gravy during storage at abuse temperatures (15 and 22 degrees C) was assessed by inoculation studies. Irradiation resulted in a 3-4 log10 reduction in numbers of both pathogens. Whenever B. cereus and S. aureus numbers reached 10(6) and 10(7) cfu/g, respectively, during storage their toxins were detectable. As the time taken to attain these levels was longer in irradiated than in unirradiated samples, toxin production by both pathogens was delayed by irradiation. When samples initially containing low levels (10(2)/g) of S. aureus were irradiated no toxin was produced during subsequent storage at 15 or 22 degrees C. Diarrhoeal toxin produced by B. cereus was detected after 2 days at 22 degrees C, but not at 15 degrees C, in samples containing 10(2) cells/g prior to irradiation. When higher numbers (10(6)/g) of either pathogen were present prior to irradiation, toxins were produced by both pathogens at 22 degrees C, but not at 15 degrees C. Microbial competition had an effect on the growth of B. cereus and S. aureus after irradiation when a low initial inoculum was applied. However, when a higher inoculum was used the pathogens outnumbered their competitors and competition effects were less important. It was concluded that low-dose irradiation would improve the microbiological safety of roast beef and gravy.

  19. High temperature in combination with UV irradiation enhances horizontal transfer of stx2 gene from E. coli O157:H7 to non-pathogenic E. coli.

    Directory of Open Access Journals (Sweden)

    Wan-Fu Yue

    Full Text Available Shiga toxin (stx genes have been transferred to numerous bacteria, one of which is E. coli O157:H7. It is a common belief that stx gene is transferred by bacteriophages, because stx genes are located on lambdoid prophages in the E. coli O157:H7 genome. Both E. coli O157:H7 and non-pathogenic E. coli are highly enriched in cattle feedlots. We hypothesized that strong UV radiation in combination with high temperature accelerates stx gene transfer into non-pathogenic E. coli in feedlots.E. coli O157:H7 EDL933 strain were subjected to different UV irradiation (0 or 0.5 kJ/m(2 combination with different temperature (22, 28, 30, 32, and 37 °C treatments, and the activation of lambdoid prophages was analyzed by plaque forming unit while induction of Stx2 prophages was quantified by quantitative real-time PCR. Data showed that lambdoid prophages in E. coli O157:H7, including phages carrying stx2, were activated under UV radiation, a process enhanced by elevated temperature. Consistently, western blotting analysis indicated that the production of Shiga toxin 2 was also dramatically increased by UV irradiation and high temperature. In situ colony hybridization screening indicated that these activated Stx2 prophages were capable of converting laboratory strain of E. coli K12 into new Shiga toxigenic E. coli, which were further confirmed by PCR and ELISA analysis.These data implicate that high environmental temperature in combination with UV irradiation accelerates the spread of stx genes through enhancing Stx prophage induction and Stx phage mediated gene transfer. Cattle feedlot sludge are teemed with E. coli O157:H7 and non-pathogenic E. coli, and is frequently exposed to UV radiation via sunlight, which may contribute to the rapid spread of stx gene to non-pathogenic E. coli and diversity of shiga toxin producing E. coli.

  20. Expression of deleted, atoxic atypical recombinant beta2 toxin in a baculovirus system and production of polyclonal and monoclonal antibodies.

    Science.gov (United States)

    Serroni, Anna; Magistrali, Chiara Francesca; Pezzotti, Giovanni; Bano, Luca; Pellegrini, Martina; Severi, Giulio; Di Pancrazio, Chiara; Luciani, Mirella; Tittarelli, Manuela; Tofani, Silvia; De Giuseppe, Antonio

    2017-05-25

    Clostridium perfringens is an important animal and human pathogen that can produce more than 16 different major and minor toxins. The beta-2 minor toxin (CPB2), comprising atypical and consensus variants, appears to be involved in both human and animal enterotoxaemia syndrome. The exact role of CPB2 in pathogenesis is poorly investigated, and its mechanism of action at the molecular level is still unknown because of the lack of specific reagents such as monoclonal antibodies against the CPB2 protein and/or the availability of a highly purified antigen. Previous studies have reported that purified wild-type or recombinant CPB2 toxin, expressed in a heterologous system, presented cytotoxic effects on human intestinal cell lines. Undoubtedly, for this reason, to date, these purified proteins have not yet been used for the production of monoclonal antibodies (MAbs). Recently, monoclonal antibodies against CPB2 were generated using peptides designed on predicted antigenic epitopes of this toxin. In this paper we report, for the first time, the expression in a baculovirus system of a deleted recombinant C-terminal 6xHis-tagged atypical CPB2 toxin (rCPB2 Δ1-25 -His 6 ) lacking the 25 amino acids (aa) of the N-terminal putative signal sequence. A high level of purified recombinant rCPB2 Δ1-25 -His 6 was obtained after purification by Ni 2+ affinity chromatography. The purified product showed no in vitro and in vivo toxicity. Polyclonal antibodies and twenty hybridoma-secreting Mabs were generated using purified rCPB2 Δ1-25 -His 6 . Finally, the reactivity and specificity of the new antibodies were tested against both recombinant and wild-type CPB2 toxins. The high-throughput of purified atoxic recombinant CPB2 produced in insect cells, allowed to obtain monoclonal and polyclonal antibodies. The availability of these molecules could contribute to develop immunoenzymatic methods and/or to perform studies about the biological activity of CPB2 toxin.

  1. Toxins of filamentous fungi.

    Science.gov (United States)

    Bhatnagar, Deepak; Yu, Jiujiang; Ehrlich, Kenneth C

    2002-01-01

    Mycotoxins are low-molecular-weight secondary metabolites of fungi. The most significant mycotoxins are contaminants of agricultural commodities, foods and feeds. Fungi that produce these toxins do so both prior to harvest and during storage. Although contamination of commodities by toxigenic fungi occurs frequently in areas with a hot and humid climate (i.e. conditions favorable for fungal growth), they can also be found in temperate conditions. Production of mycotoxins is dependent upon the type of producing fungus and environmental conditions such as the substrate, water activity (moisture and relative humidity), duration of exposure to stress conditions and microbial, insect or other animal interactions. Although outbreaks of mycotoxicoses in humans have been documented, several of these have not been well characterized, neither has a direct correlation between the mycotoxin and resulting toxic effect been well established in vivo. Even though the specific modes of action of most of the toxins are not well established, acute and chronic effects in prokaryotic and eukaryotic systems, including humans have been reported. The toxicity of the mycotoxins varies considerably with the toxin, the animal species exposed to it, and the extent of exposure, age and nutritional status. Most of the toxic effects of mycotoxins are limited to specific organs, but several mycotoxins affect many organs. Induction of cancer by some mycotoxins is a major concern as a chronic effect of these toxins. It is nearly impossible to eliminate mycotoxins from the foods and feed in spite of the regulatory efforts at the national and international levels to remove the contaminated commodities. This is because mycotoxins are highly stable compounds, the producing fungi are ubiquitous, and food contamination can occur both before and after harvest. Nevertheless, good farm management practices and adequate storage facilities minimize the toxin contamination problems. Current research is

  2. Dinophysis Toxins: Causative Organisms, Distribution and Fate in Shellfish

    Directory of Open Access Journals (Sweden)

    Beatriz Reguera

    2014-01-01

    Full Text Available Several Dinophysis species produce diarrhoetic toxins (okadaic acid and dinophysistoxins and pectenotoxins, and cause gastointestinal illness, Diarrhetic Shellfish Poisoning (DSP, even at low cell densities (<103 cells·L−1. They are the main threat, in terms of days of harvesting bans, to aquaculture in Northern Japan, Chile, and Europe. Toxicity and toxin profiles are very variable, more between strains than species. The distribution of DSP events mirrors that of shellfish production areas that have implemented toxin regulations, otherwise misinterpreted as bacterial or viral contamination. Field observations and laboratory experiments have shown that most of the toxins produced by Dinophysis are released into the medium, raising questions about the ecological role of extracelular toxins and their potential uptake by shellfish. Shellfish contamination results from a complex balance between food selection, adsorption, species-specific enzymatic transformations, and allometric processes. Highest risk areas are those combining Dinophysis strains with high cell content of okadaates, aquaculture with predominance of mytilids (good accumulators of toxins, and consumers who frequently include mussels in their diet. Regions including pectenotoxins in their regulated phycotoxins will suffer from much longer harvesting bans and from disloyal competition with production areas where these toxins have been deregulated.

  3. Update on botulinum toxin and dermal fillers.

    Science.gov (United States)

    Berbos, Zachary J; Lipham, William J

    2010-09-01

    The art and science of facial rejuvenation is an ever-evolving field of medicine, as evidenced by the continual development of new surgical and nonsurgical treatment modalities. Over the past 10 years, the use of botulinum toxin and dermal fillers for aesthetic purposes has risen sharply. Herein, we discuss properties of several commonly used injectable products and provide basic instruction for their use toward the goal of achieving facial rejuvenation. The demand for nonsurgical injection-based facial rejuvenation products has risen enormously in recent years. Used independently or concurrently, botulinum toxin and dermal filler agents offer an affordable, minimally invasive approach to facial rejuvenation. Botulinum toxin and dermal fillers can be used to diminish facial rhytides, restore facial volume, and sculpt facial contours, thereby achieving an aesthetically pleasing, youthful facial appearance.

  4. The Regulatory Networks That Control Clostridium difficile Toxin Synthesis

    Science.gov (United States)

    Martin-Verstraete, Isabelle; Peltier, Johann; Dupuy, Bruno

    2016-01-01

    The pathogenic clostridia cause many human and animal diseases, which typically arise as a consequence of the production of potent exotoxins. Among the enterotoxic clostridia, Clostridium difficile is the main causative agent of nosocomial intestinal infections in adults with a compromised gut microbiota caused by antibiotic treatment. The symptoms of C. difficile infection are essentially caused by the production of two exotoxins: TcdA and TcdB. Moreover, for severe forms of disease, the spectrum of diseases caused by C. difficile has also been correlated to the levels of toxins that are produced during host infection. This observation strengthened the idea that the regulation of toxin synthesis is an important part of C. difficile pathogenesis. This review summarizes our current knowledge about the regulators and sigma factors that have been reported to control toxin gene expression in response to several environmental signals and stresses, including the availability of certain carbon sources and amino acids, or to signaling molecules, such as the autoinducing peptides of quorum sensing systems. The overlapping regulation of key metabolic pathways and toxin synthesis strongly suggests that toxin production is a complex response that is triggered by bacteria in response to particular states of nutrient availability during infection. PMID:27187475

  5. Special issue: engineering toxins for 21st-century therapies: introduction.

    Science.gov (United States)

    Acharya, K Ravi

    2011-12-01

    This special issue on 'Engineering toxins for 21st century therapies' provides a critical review of the current state of multifaceted aspects of toxin research by some of the leading researchers in the field. It also highlights the clinical potential and challenges for development of novel biologics based on engineered toxin derived products. © 2011 The Author Journal compilation © 2011 FEBS.

  6. Stool C difficile toxin

    Science.gov (United States)

    ... toxin; Colitis - toxin; Pseudomembranous - toxin; Necrotizing colitis - toxin; C difficile - toxin ... be analyzed. There are several ways to detect C difficile toxin in the stool sample. Enzyme immunoassay ( ...

  7. Shiga toxin-producing Escherichia coli

    DEFF Research Database (Denmark)

    Pedersen, Rune Micha; Nielsen, Marc Trunjer Kusk; Möller, Sören

    2018-01-01

    samples of patients with suspected infective gastroenteritis were analysed for STEC. METHODS: In this population-based cohort study, all stool samples referred to two clinical microbiology laboratories, were screened for STEC by culture and/or PCR. Epidemiological (n=170) and clinical (n=209...

  8. Alternaria Toxins: Potential Virulence Factors and Genes Related to Pathogenesis

    Directory of Open Access Journals (Sweden)

    Mukesh Meena

    2017-08-01

    Full Text Available Alternaria is an important fungus to study due to their different life style from saprophytes to endophytes and a very successful fungal pathogen that causes diseases to a number of economically important crops. Alternaria species have been well-characterized for the production of different host-specific toxins (HSTs and non-host specific toxins (nHSTs which depend upon their physiological and morphological stages. The pathogenicity of Alternaria species depends on host susceptibility or resistance as well as quantitative production of HSTs and nHSTs. These toxins are chemically low molecular weight secondary metabolites (SMs. The effects of toxins are mainly on different parts of cells like mitochondria, chloroplast, plasma membrane, Golgi complex, nucleus, etc. Alternaria species produce several nHSTs such as brefeldin A, tenuazonic acid, tentoxin, and zinniol. HSTs that act in very low concentrations affect only certain plant varieties or genotype and play a role in determining the host range of specificity of plant pathogens. The commonly known HSTs are AAL-, AK-, AM-, AF-, ACR-, and ACT-toxins which are named by their host specificity and these toxins are classified into different family groups. The HSTs are differentiated on the basis of bio-statistical and other molecular analyses. All these toxins have different mode of action, biochemical reactions and signaling mechanisms to cause diseases. Different species of Alternaria produced toxins which reveal its biochemical and genetic effects on itself as well as on its host cells tissues. The genes responsible for the production of HSTs are found on the conditionally dispensable chromosomes (CDCs which have been well characterized. Different bio-statistical methods like basic local alignment search tool (BLAST data analysis used for the annotation of gene prediction, pathogenicity-related genes may provide surprising knowledge in present and future.

  9. Effect of sodium nitrite on toxin production by Clostridium botulinum in bacon.

    Science.gov (United States)

    Christiansen, L N; Tompkin, R B; Shaparis, A B; Kueper, T V; Johnston, R W; Kautter, D A; Kolari, O J

    1974-04-01

    Pork bellies were formulated to 0, 30, 60, 120, 170, or 340 mug of nitrite per g of meat and inoculated with Clostridium botulinum via pickle or after processing and slicing. Processed bacon was stored at 7 or 27 C and assayed for nitrite, nitrate, and botulinal toxin at different intervals. Nitrite levels declined during processing and storage. The rate of decrease was more rapid at 27 than at 7 C. Although not added to the system, nitrate was detected in samples during processing and storage at 7 and 27 C. The amount of nitrate found was related to formulated nitrite levels. No toxin was found in samples incubated at 7 C throughout the 84-day test period. At 27 C, via pickle, inoculated samples with low inoculum (210 C. botulinum per g before processing and 52 per g after processing) became toxic if formulated with 120 mug of nitrite per g of meat or less. Toxin was not detected in bacon formulated with 170 or 340 mug of nitrite per g of meat under these same conditions. Toxin was detected at all formulated nitrite levels in bacon inoculated via the pickle with 19,000 C. botulinum per g (4,300 per g after processing) and in samples inoculated after slicing. However, increased levels of formulated nitrite decreased the probability of botulinal toxin formation in bacon inoculated by both methods.

  10. Characterizing relationships among fecal indicator bacteria ...

    Science.gov (United States)

    Bed sediments of streams and rivers may store high concentrations of fecal indicator bacteria (FIB) and pathogens. Due to resuspension events, these contaminants can be mobilized into the water column and affect overall water quality. Other bacterial indicators such as microbial source tracking (MST) markers, developed to determine potential sources of fecal contamination, can also be resuspended from bed sediments. The primary objective of this study was to predict occurrence of waterborne pathogens in water and streambed sediments using a simple statistical model that includes traditionally measured FIB, environmental parameters and source allocation, using MST markers as predictor variables. Synoptic sampling events were conducted during baseflow conditions downstream from agricultural (AG), forested (FORS), and wastewater pollution control plant (WPCP) land uses. Concentrations of FIB and MST markers were measured in water and sediments, along with occurrences of the enteric pathogens Campylobacter, Listeria and Salmonella, and the virulence gene that carries Shiga toxin, stx2. Pathogens were detected in water more often than in underlying sediments. Shiga toxin was significantly related to land use, with concentrations of the ruminant marker selected as an independent variable that could correctly classify 76% and 64% of observed Shiga toxin occurrences in water and sediment, respectively. FIB concentrations and water quality parameters were also selected a

  11. Two enzymes involved in biosynthesis of the host-selective phytotoxin HC-toxin

    International Nuclear Information System (INIS)

    Walton, J.D.

    1987-01-01

    Cochliobolus carbonum race 1 produces a cyclic tetrapeptide HC-toxin, which is necessary for its exceptional virulence on certain varieties of maize. Previous genetic analysis of HC-toxin production by the fungus has indicated that a single genetic locus controls HC-toxin production. Enzymes involved in the biosynthesis of HC-toxin have been sought by following the precedents established for the biosynthetic enzymes of cyclic peptide antibiotics. Two enzymatic activities from C. carbonum race 1 were found, a D-alanine- and an L-proline-dependent ATP/PP/sub i/ exchange, which by biochemical and genetic criteria were shown to be involved in the biosynthesis of HC-toxin. These two activities were present in all tested race 1 isolates of C. carbonum, which produce HC-toxin, and in none of the tested race 2 and race 3 isolates, which do not produce the toxin. In a genetic cross between two isolates of C. carbonum differing at the tox locus, all tox + progeny had both activities, and all tox - progeny lacked both activities

  12. Structural Basis for the Specific Neutralization of Stx2a with a Camelid Single Domain Antibody Fragment

    Directory of Open Access Journals (Sweden)

    Robert Alvin Bernedo-Navarro

    2018-03-01

    Full Text Available Background: Shiga toxin-producing Escherichia coli (STEC are a subset of pathogens leading to illnesses such as diarrhea, hemolytic uremic syndrome and even death. The Shiga toxins are the main virulence factors and divided in two groups: Stx1 and Stx2, of which the latter is more frequently associated with severe pathologies in humans. Results: An immune library of nanobodies (Nbs was constructed after immunizing an alpaca with recombinant Shiga toxin-2a B subunit (rStx2aB, to retrieve multiple rStx2aB-specific Nbs. The specificity of five Nbs towards rStx2aB was confirmed in ELISA and Western blot. Nb113 had the highest affinity (9.6 nM and its bivalent construct exhibited a 100-fold higher functional affinity. The structure of the Nb113 in complex with rStx2aB was determined via X-ray crystallography. The crystal structure of the Nb113–rStx2aB complex revealed that five copies of Nb113 bind to the rStx2aB pentamer and that the Nb113 epitope overlaps with the Gb3 binding site, thereby providing a structural basis for the neutralization of Stx2a by Nb113 that was observed on Vero cells. Finally, the tandem-repeated, bivalent Nb1132 exhibits a higher toxin neutralization capacity compared to monovalent Nb113. Conclusions: The Nb of highest affinity for rStx2aB is also the best Stx2a and Stx2c toxin neutralizing Nb, especially in a bivalent format. This lead Nb neutralizes Stx2a by competing for the Gb3 receptor. The fusion of the bivalent Nb1132 with a serum albumin specific Nb is expected to combine high toxin neutralization potential with prolonged blood circulation.

  13. BiP Negatively Affects Ricin Transport

    Directory of Open Access Journals (Sweden)

    Kirsten Sandvig

    2013-05-01

    Full Text Available The AB plant toxin ricin binds both glycoproteins and glycolipids at the cell surface via its B subunit. After binding, ricin is endocytosed and then transported retrogradely through the Golgi to the endoplasmic reticulum (ER. In the ER, the A subunit is retrotranslocated to the cytosol in a chaperone-dependent process, which is not fully explored. Recently two separate siRNA screens have demonstrated that ER chaperones have implications for ricin toxicity. ER associated degradation (ERAD involves translocation of misfolded proteins from ER to cytosol and it is conceivable that protein toxins exploit this pathway. The ER chaperone BiP is an important ER regulator and has been implicated in toxicity mediated by cholera and Shiga toxin. In this study, we have investigated the role of BiP in ricin translocation to the cytosol. We first show that overexpression of BiP inhibited ricin translocation and protected cells against the toxin. Furthermore, shRNA-mediated depletion of BiP enhanced toxin translocation resulting in increased cytotoxicity. BiP-dependent inhibition of ricin toxicity was independent of ER stress. Our findings suggest that in contrast to what was shown with the Shiga toxin, the presence of BiP does not facilitate, but rather inhibits the entry of ricin into the cytosol.

  14. 9 CFR 121.3 - VS select agents and toxins.

    Science.gov (United States)

    2010-01-01

    ... genetically modified. (d) VS select agents or toxins that meet any of the following criteria are excluded from... AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS POSSESSION, USE, AND... recombinant organisms: (1) Nucleic acids that can produce infectious forms of any of the select agent viruses...

  15. Evaluation of Cholera Toxin Expression in Acidic, Alkaline and Neutral Conditions

    Directory of Open Access Journals (Sweden)

    Narges Rahimi

    2015-02-01

    Full Text Available Background: Cholera is a severe disease which is caused by Vibrio cholerae and it is typically transmitted by either contaminated food or water particularly in developing countries. The most important virulence factor of this bacterium is an enterotoxin called cholera toxin which is a protein complex secreted by the Vibrio cholerae. Objectives: In this project, we determined the production of cholera toxin at different pH values. Materials and Methods: Two standard strain of Vibrio cholerae O1 biovar EL Tor N16961 and Vibrio cholerae O1 biovar Classic ATCC 14035 were used. After overnight cultivation of both the strains the total mRNA extracted and converted to total cDNA. Results: By Relative Real-Time PCR analysis the most cholera toxin production in classical and El Tor strains was at pH 8.5 and 8, respectively. Conclusions: Therefore, We may conclude that use of acidic diet will help in reduction of cholera toxin production.

  16. Inhibition of cholera toxin and other AB toxins by polyphenolic compounds

    Science.gov (United States)

    All AB-type protein toxins have intracellular targets despite an initial extracellular location. These toxins use different methods to reach the cytosol and have different effects on the target cell. Broad-spectrum inhibitors against AB toxins are therefore hard to develop because the toxins use dif...

  17. Acid Sphingomyelinase Promotes Cellular Internalization of Clostridium perfringens Iota-Toxin.

    Science.gov (United States)

    Nagahama, Masahiro; Takehara, Masaya; Miyamoto, Kazuaki; Ishidoh, Kazumi; Kobayashi, Keiko

    2018-05-20

    Clostridium perfringens iota-toxin is a binary actin-ADP-ribosylating toxin composed of the enzymatic component Ia and receptor binding component Ib. Ib binds to a cell surface receptor, forms Ib oligomer in lipid rafts, and associates with Ia. The Ia-Ib complex then internalizes by endocytosis. Here, we showed that acid sphingomyelinase (ASMase) facilitates the cellular uptake of iota-toxin. Inhibitions of ASMase and lysosomal exocytosis by respective blockers depressed cell rounding induced by iota-toxin. The cytotoxicity of the toxin increased in the presence of Ca 2+ in extracellular fluids. Ib entered target cells in the presence but not the absence of Ca 2+ . Ib induced the extracellular release of ASMase in the presence of Ca 2+ . ASMase siRNA prevented the cell rounding induced by iota-toxin. Furthermore, treatment of the cells with Ib resulted in the production of ceramide in cytoplasmic vesicles. These observations showed that ASMase promotes the internalization of iota-toxin into target cells.

  18. Particle size fractionation of paralytic shellfish toxins (PSTs): seasonal distribution and bacterial production in the St Lawrence estuary, Canada.

    Science.gov (United States)

    Michaud, S; Levasseur, M; Doucette, G; Cantin, G

    2002-10-01

    We determined the seasonal distribution of paralytic shellfish toxins (PSTs) and PST producing bacteria in > 15, 5-15, and 0.22-5 microm size fractions in the St Lawrence. We also measured PSTs in a local population of Mytilus edulis. PST concentrations were determined in each size fraction and in laboratory incubations of sub-samples by high performance liquid chromatography (HPLC), including the rigorous elimination of suspected toxin 'imposter' peaks. Mussel toxin levels were determined by mouse bioassay and HPLC. PSTs were detected in all size fractions during the summer sampling season, with 47% of the water column toxin levels associated with particles smaller than Alexandrium tamarense ( 15 microm size fraction, we estimated that as much as 92% of PSTs could be associated with particles other than A. tamarense. Our results stress the importance of taking into account the potential presence of PSTs in size fractions other than that containing the known algal producer when attempting to model shellfish intoxication, especially during years of low cell abundance. Finally, our HPLC results confirmed the presence of bacteria capable of autonomous PST production in the St Lawrence as well as demonstrating their regular presence and apparent diversity in the plankton. Copyright 2002 Elsevier Science Ltd.

  19. Targeting Staphylococcus aureus Toxins: A Potential form of Anti-Virulence Therapy

    Directory of Open Access Journals (Sweden)

    Cin Kong

    2016-03-01

    Full Text Available Staphylococcus aureus is an opportunistic pathogen and the leading cause of a wide range of severe clinical infections. The range of diseases reflects the diversity of virulence factors produced by this pathogen. To establish an infection in the host, S. aureus expresses an inclusive set of virulence factors such as toxins, enzymes, adhesins, and other surface proteins that allow the pathogen to survive under extreme conditions and are essential for the bacteria’s ability to spread through tissues. Expression and secretion of this array of toxins and enzymes are tightly controlled by a number of regulatory systems. S. aureus is also notorious for its ability to resist the arsenal of currently available antibiotics and dissemination of various multidrug-resistant S. aureus clones limits therapeutic options for a S. aureus infection. Recently, the development of anti-virulence therapeutics that neutralize S. aureus toxins or block the pathways that regulate toxin production has shown potential in thwarting the bacteria’s acquisition of antibiotic resistance. In this review, we provide insights into the regulation of S. aureus toxin production and potential anti-virulence strategies that target S. aureus toxins.

  20. Binding of ATP by pertussis toxin and isolated toxin subunits

    International Nuclear Information System (INIS)

    Hausman, S.Z.; Manclark, C.R.; Burns, D.L.

    1990-01-01

    The binding of ATP to pertussis toxin and its components, the A subunit and B oligomer, was investigated. Whereas, radiolabeled ATP bound to the B oligomer and pertussis toxin, no binding to the A subunit was observed. The binding of [ 3 H]ATP to pertussis toxin and the B oligomer was inhibited by nucleotides. The relative effectiveness of the nucleotides was shown to be ATP > GTP > CTP > TTP for pertussis toxin and ATP > GTP > TTP > CTP for the B oligomer. Phosphate ions inhibited the binding of [ 3 H]ATP to pertussis toxin in a competitive manner; however, the presence of phosphate ions was essential for binding of ATP to the B oligomer. The toxin substrate, NAD, did not affect the binding of [ 3 H]ATP to pertussis toxin, although the glycoprotein fetuin significantly decreased binding. These results suggest that the binding site for ATP is located on the B oligomer and is distinct from the enzymatically active site but may be located near the eukaryotic receptor binding site

  1. Binding of ATP by pertussis toxin and isolated toxin subunits

    Energy Technology Data Exchange (ETDEWEB)

    Hausman, S.Z.; Manclark, C.R.; Burns, D.L. (Center for Biologics Evaluation and Research, Bethesda, MD (USA))

    1990-07-03

    The binding of ATP to pertussis toxin and its components, the A subunit and B oligomer, was investigated. Whereas, radiolabeled ATP bound to the B oligomer and pertussis toxin, no binding to the A subunit was observed. The binding of ({sup 3}H)ATP to pertussis toxin and the B oligomer was inhibited by nucleotides. The relative effectiveness of the nucleotides was shown to be ATP > GTP > CTP > TTP for pertussis toxin and ATP > GTP > TTP > CTP for the B oligomer. Phosphate ions inhibited the binding of ({sup 3}H)ATP to pertussis toxin in a competitive manner; however, the presence of phosphate ions was essential for binding of ATP to the B oligomer. The toxin substrate, NAD, did not affect the binding of ({sup 3}H)ATP to pertussis toxin, although the glycoprotein fetuin significantly decreased binding. These results suggest that the binding site for ATP is located on the B oligomer and is distinct from the enzymatically active site but may be located near the eukaryotic receptor binding site.

  2. Mutations in the Histone-like Nucleoid Structuring Regulatory Gene (hns) Decrease the Adherence of Shiga Toxin-producing Escherichia coli 091:H21 Strain B2F1 to Human Colonic Epithelial Cells and Increase the Production of Hemolysin

    Science.gov (United States)

    1999-10-19

    osmoregulation of outer membrane proteins and virulence determinants in Vibrio cholerae requires toxR. J. Bacteriol. 170:2575-2583. Mobley, H. L., D. M. Green...produced by ETEC organisms is homologous to the toxin encoded by Y: cholerae . These toxins are the primary cause of the watery diarrhea associated with ETEC...Escherichia coli as a cause ofdiarrhea among children in Mexico . J. Clin. Microbiol. 25:1913-1919. Maurelli, A. T., and P. J. Sansonetti. 1988

  3. T-2 toxin Analysis in Poultry and Cattle Feedstuff.

    Science.gov (United States)

    Gholampour Azizi, Issa; Azarmi, Masumeh; Danesh Pouya, Naser; Rouhi, Samaneh

    2014-05-01

    T-2 toxin is a mycotoxin that is produced by the Fusarium fungi. Consumption of food and feed contaminated with T-2 toxin causes diseases in humans and animals. In this study T-2 toxin was analyzed in poultry and cattle feedstuff in cities of Mazandaran province (Babol, Sari, Chalus), Northern Iran. In this study, 90 samples were analyzed for T-2 toxin contamination by the ELISA method. Out of 60 concentrate and bagasse samples collected from various cities of Mazandaran province, 11.7% and 3.3% were contaminated with T-2 toxin at concentrations > 25 and 50 µg/kg, respectively. For mixed poultry diets, while 10% of the 30 analyzed samples were contaminated with > 25 µg/kg, none of the tested samples contained T-2 toxin at levels > 50 µg/kg. The results obtained from this study show that poultry and cattle feedstuff can be contaminated with different amounts of T-2 toxin in different conditions and locations. Feedstuff that are contaminated by this toxin cause different diseases in animals; thus, potential transfer of mycotoxins to edible by-products from animals fed mycotoxin-contaminated feeds drives the need to routinely monitor mycotoxins in animal feeds and their components. This is the basis on which effective management of mycotoxins and their effects can be implemented.

  4. Effects of Phenolic Acids on the Growth and Production of T-2 and HT-2 Toxins by Fusarium langsethiae and F. sporotrichioides.

    Science.gov (United States)

    Ferruz, Elena; Atanasova-Pénichon, Vessela; Bonnin-Verdal, Marie-Noëlle; Marchegay, Gisèle; Pinson-Gadais, Laëtitia; Ducos, Christine; Lorán, Susana; Ariño, Agustín; Barreau, Christian; Richard-Forget, Florence

    2016-04-04

    The effect of natural phenolic acids was tested on the growth and production of T-2 and HT-2 toxins by Fusarium langsethiae and F. sporotrichioides, on Mycotoxin Synthetic medium. Plates treated with 0.5 mM of each phenolic acid (caffeic, chlorogenic, ferulic and p-coumaric) and controls without phenolic acid were incubated for 14 days at 25 °C. Fungal biomass of F. langsethiae and F. sporotrichioides was not reduced by the phenolic acids. However, biosynthesis of T-2 toxin by F. langsethiae was significantly reduced by chlorogenic (23.1%) and ferulic (26.5%) acids. Production of T-2 by F. sporotrichioides also decreased with ferulic acid by 23% (p phenolic acids had a variable effect on fungal growth and mycotoxin production, depending on the strain and the concentration and type of phenolic acid assayed.

  5. Failure of botulinum toxin injection for neurogenic detrusor overactivity: Switch of toxin versus second injection of the same toxin.

    Science.gov (United States)

    Peyronnet, Benoit; Castel-Lacanal, Evelyne; Manunta, Andréa; Roumiguié, Mathieu; Marque, Philippe; Rischmann, Pascal; Gamé, Xavier

    2015-12-01

    To evaluate the efficacy of a second injection of the same toxin versus switching to a different botulinum toxin A after failure of a first detrusor injection in patients with neurogenic detrusor overactivity. The charts of all patients who underwent detrusor injections of botulinum toxin A (either abobotulinumtoxinA or onabotulinumtoxinA) for the management of neurogenic detrusor overactivity at a single institution were retrospectively reviewed. Patients in whom a first detrusor injection had failed were included in the present study. They were managed by a second injection of the same toxin at the same dosage or by a new detrusor injection using a different botulinum toxin A. Success was defined as a resolution of urgency, urinary incontinence and detrusor overactivity in a patient self-catheterizing seven times or less per 24 h. A total of 58 patients were included for analysis. A toxin switch was carried out in 29 patients, whereas the other 29 patients received a reinjection of the same toxin at the same dose. The success rate was higher in patients who received a toxin switch (51.7% vs. 24.1%, P = 0.03). Patients treated with a switch from abobotulinumtoxinA to onabotulinumtoxinA and those treated with a switch from onabotulinumtoxinA to abobotulinumtoxinA had similar success rates (52.9% vs. 50%, P = 0.88). After failure of a first detrusor injection of botulinum toxin for neurogenic detrusor overactivity, a switch to a different toxin seems to be more effective than a second injection of the same toxin. The replacement of onabotulinumtoxin by abobotulinumtoxin or the reverse provides similar results. © 2015 The Japanese Urological Association.

  6. Radiolabelling of cholera toxin

    International Nuclear Information System (INIS)

    Santos, R.G.; Neves, Nicoli M.J.; Abdalla, L.F.; Brandao, R.L.; Etchehebehere, L.; Lima, M.E. de; Nicoli, J.R.

    1999-01-01

    Binding of cholera toxin to ganglioside receptors of enterocyte microvilli catalyzes the activation of adenylate cyclase causing a rise in cAMP which final result is a copious diarrhea. Saccharomyces boulardii, a nonpathogenic yeast has been used to prevent diarrhea. Although the antidiarrheic properties of S. boulardii are widely recognized, this yeast has been used on empirical basis, and the mechanism of this protective effect is unknown. The addition of cholera toxin to S. boulardii induces the raising of cAMP that triggers the activation of neutral trehalase. This suggests that toxin specifically binding to cells, is internalized and active the protein phosphorylation cascade. Our objective is labeling the cholera toxin to verify the presence of binding sites on yeast cell surfaces for the cholera toxin. Cholera toxin was radiolabelled with Na 125 I by a chloramine-T method modified from Cuatrecasas and Griffiths et alii. The 125 I-Cholera toxin showed a specific radioactivity at about 1000 cpm/fmol toxin. Biological activity of labeled cholera toxin measured by trehalase activation was similar to the native toxin. (author)

  7. Milling technological experiments to reduce Fusarium toxin contamination in wheat

    Directory of Open Access Journals (Sweden)

    Véha A.

    2015-01-01

    Full Text Available We examine 4 different DON-toxin-containing (0.74 - 1.15 - 1.19 - 2.14 mg/kg winter wheat samples: they were debranned and undebranned, and we investigated the flour’s and the by-products’ (coarse, fine bran toxin content changes. SATAKE lab-debranner was used for debranning and BRABENDER lab-mill for the milling process. Without debranning, two sample flours were above the DON toxin limit (0.75 mg/kg, which are waste. By minimum debranning (and minimum debranning mass loss; 6-8%, our experience with whole flour is that the multi-stage debranning measurement significantly reduces the content of the flour’s DON toxin, while the milling by-products, only after careful consideration and DON toxin measurements, may be produced for public consumption and for feeding.

  8. T-2 toxin and HT-2 toxin in grain and grain-based commodities in Europe: occurrence, factors affecting occurrence, co-occurence and toxicological effects

    NARCIS (Netherlands)

    Fels-Klerx, van der H.J.; Stratakou, I.

    2010-01-01

    This paper presents an overview of the occurrence of T-2 toxin and HT-2 toxin in cereals in Europe and derived food products, factors influencing the occurrence, co-occurrence with other trichothecenes, and toxicological effects of T-2 and HT-2 in human. Of all cereals, oats showed to be most

  9. Updates on tetanus toxin: a fundamental approach

    Directory of Open Access Journals (Sweden)

    Md. Ahaduzzaman

    2015-03-01

    Full Text Available Clostridium tetani is an anaerobic bacterium that produces second most poisonous protein toxins than any other bacteria. Tetanus in animals is sporadic in nature but difficult to combat even by using antibiotics and antiserum. It is crucial to understand the fundamental mechanisms and signals that control toxin production for advance research and medicinal uses. This review was intended for better understanding the basic patho-physiology of tetanus and neurotoxins (TeNT among the audience of related field.

  10. Multiplex polymerase chain reaction (PCR) assay for simultaneous ...

    African Journals Online (AJOL)

    user

    2011-02-28

    Feb 28, 2011 ... To evaluate the etiology of shiga toxin-producing E. coli (STEC), enteropathogenic E. ... National Center for Biotechnology Information; BLAST, Basic ... Poor sanitation, personal ..... Mem Inst Oswaldo Cruz, Rio de Janeiro.

  11. Radiolabelling of cholera toxin

    Energy Technology Data Exchange (ETDEWEB)

    Santos, R.G.; Neves, Nicoli M.J. [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN), Belo Horizonte, MG (Brazil); Abdalla, L.F.; Brandao, R.L.; Etchehebehere, L. [Ouro Preto Univ., MG (Brazil). Escola de Farmacia. Lab. de Fisiologia e Bioquimica de Microorganismos; Lima, M.E. de [Minas Gerais Univ., Belo Horizonte, MG (Brazil). Inst. de Ciencias Biologicas. Dept. de Bioquimica e Imunologia; Nicoli, J.R. [Minas Gerais Univ., Belo Horizonte, MG (Brazil). Inst. de Ciencias Biologicas. Dept. de Microbiologia

    1999-11-01

    Binding of cholera toxin to ganglioside receptors of enterocyte microvilli catalyzes the activation of adenylate cyclase causing a rise in cAMP which final result is a copious diarrhea. Saccharomyces boulardii, a nonpathogenic yeast has been used to prevent diarrhea. Although the antidiarrheic properties of S. boulardii are widely recognized, this yeast has been used on empirical basis, and the mechanism of this protective effect is unknown. The addition of cholera toxin to S. boulardii induces the raising of cAMP that triggers the activation of neutral trehalase. This suggests that toxin specifically binding to cells, is internalized and active the protein phosphorylation cascade. Our objective is labeling the cholera toxin to verify the presence of binding sites on yeast cell surfaces for the cholera toxin. Cholera toxin was radiolabelled with Na {sup 125} I by a chloramine-T method modified from Cuatrecasas and Griffiths et alii. The {sup 125} I-Cholera toxin showed a specific radioactivity at about 1000 cpm/fmol toxin. Biological activity of labeled cholera toxin measured by trehalase activation was similar to the native toxin. (author) 5 refs., 3 figs.; e-mail: nevesmj at urano.cdtn.br

  12. Use of an Electrostatic Spraying System or the Sprayed Lethality in Container Method To Deliver Antimicrobial Agents onto the Surface of Beef Subprimals To Control Shiga Toxin-Producing Escherichia coli.

    Science.gov (United States)

    Stella, J Max; Luchansky, John B; Miller, Kelsey; Shoyer, Bradley A; Shane, Laura E; McGeary, Lianna; Osoria, Manuela; Stahler, Laura J; Sevart, Nicholas J; Phebus, Randall K; Thippareddi, Harshavardhan; Porto-Fett, Anna C S

    2017-08-01

    The efficacy of an electrostatic spraying system (ESS) and/or the sprayed lethality in container (SLIC) method to deliver antimicrobial agents onto the surface of beef subprimals to reduce levels of Shiga toxin-producing Escherichia coli (STEC) was evaluated. Beef subprimals were surface inoculated (lean side; ca. 5.8 log CFU per subprimal) with 2 mL of an eight-strain cocktail comprising single strains of rifampin-resistant (100 μg/mL) STEC (O26:H11, O45:H2, O103:H2, O104:H4, O111:H - , O121:H19, O145:NM, and O157:H7). Next, inoculated subprimals were surface treated with lauric arginate (LAE; 1%), peroxyacetic acid (PAA; 0.025%), or cetylpyridinium chloride (CPC; 0.4%) by passing each subprimal, with the inoculated lean side facing upward, through an ESS cabinet or via SLIC. Subprimals were then vacuum packaged and stored at 4°C. One set of subprimals was sampled after an additional 2 h, 3 days, or 7 days of refrigerated storage, whereas another set was retreated via SLIC after 3 days of storage with a different one of the three antimicrobial agents (e.g., a subprimal treated with LAE on day 0 was then treated with PAA or CPE on day 3). Retreated subprimals were sampled after 2 h or 4 days of additional storage at 4°C. A single initial application of LAE, PAA, or CPC via ESS or SLIC resulted in STEC reductions of ca. 0.3 to 1.3 log CFU per subprimal after 7 days of storage. However, when subprimals were initially treated with LAE, PAA, or CPC via ESS or SLIC and then separately retreated with a different one of these antimicrobial agents via SLIC on day 3, additional STEC reductions of 0.4 to 1.0 log CFU per subprimal were observed after an additional 4 days of storage. Application of LAE, PAA, or CPC, either alone or in combination, via ESS or SLIC is effective for reducing low levels (ca. 0.3 to 1.6 log CFU) of STEC that may be naturally present on the surface of beef subprimals.

  13. Consensus panel's assessment and recommendations on the use of 3 botulinum toxin type A products in facial aesthetics.

    Science.gov (United States)

    Lorenc, Z Paul; Kenkel, Jeffrey M; Fagien, Steven; Hirmand, Haideh; Nestor, Mark S; Sclafani, Anthony P; Sykes, Jonathan M; Waldorf, Heidi A

    2013-03-01

    In this summary article, the authors discuss the characteristics of abobotulinumtoxinA, incobotulinumtoxinA, and onabotulinumtoxinA. With 3 neuromodulators available in the US market, comparisons between and among products will invariably be made, so arguments for the most effective facial aesthetic uses of each neuromodulator are presented. Topics addressed in this article include patient expectations, toxin reconstitution and preparation, patient positioning, differences among products, the role of complexing proteins, and dosing and injection strategies. Recommendations are also provided by treatment area.

  14. [Questionnaire survey on medical care for male urethritis in community clinics in Shiga prefecture].

    Science.gov (United States)

    Yamashita, Hiroto; Araki, Isao; Kageyama, Susumu; Baba, Masato; Nakano, Etsuji; Okada, Yusaku

    2014-01-01

    Six regional medical associations in Shiga prefecture agreed to cooperate in an investigation of medical care for male gonococcal and chlamydial urethritis. In June 2011, we sent a questionnaire to 372 medical offices in Shiga prefecture, and analyzed replies of respondents. Ten urologists and 175 non-urologists responded to the survey (response rate 49.7%). Among 185 physicians, 52 (10 urologists and 42 nonurologists) have treated male patients with gonococcal and chlamydial urethritis. More than 20% (42/175) of non-urological clinics are involved in the medical management. At initial diagnosis for sexually transmitted male urethritis, all urologists select the nucleic acid amplification method (100%), whereas many non-urologists do not (35%). For the treatment of chlamydial urethritis, non-urologists select levofloxacin (LVFX, 52.8%) rather than azithromycin (AZM, 22.0%), whereas urologists use AZM (78.0%) mostly but do not use LVFX (0%) (p = 0.023). For the treatment of gonococcal urethritis, non-urologists prefer oral new quinolones (53.1%) compared to urologists (25.0%) (p = 0. 74). For cure judgment of gonoccocal and chlamydial urethritis, many non-urologists rely on the improvement of subjective symptoms (50 and 47%), but urologists do not (10 and 0%) (p = 0.022 and 0.026, respectively). As for recognition of the clinical guideline for sexually transmitted disease, most urologists (90%) know it, but few non-urologists (13%) do (p urethritis in Shiga prefecture. It is important to standardize the medical care for sexually transmitted male urethritis by familiarizing non-urological practitioners with the clinical guideline.

  15. Epiphytic survival of Shiga-toxigenic Escherichia coli O145 on baby spinach plants

    Science.gov (United States)

    Introduction: Non-O157 Shiga-toxigenic E. coli (STEC) have contaminated leafy greens. In early 2010, 26 confirmed illnesses were traced to a multi-state outbreak involving shredded romaine lettuce contaminated with E. coli O145, a STEC. It is possible that the O145 serotype may be suited for gro...

  16. The response of foodborne pathogens to osmotic and desiccation stresses in the food chain

    DEFF Research Database (Denmark)

    Burgess, Catherine M.; Gianotti, Andrea; Gruzdev, Nadia

    2016-01-01

    In combination with other strategies, hyperosmolarity and desiccation are frequently used by the food processing industry as a means to prevent bacterial proliferation, and particularly that of foodborne pathogens, in food products. However, it is increasingly observed that bacteria, including...... human pathogens, encode mechanisms to survive and withstand these stresses. This review provides an overview of the mechanisms employed by Salmonella spp., Shiga toxin producing E. coli, Cronobacter spp., Listeria monocytogenes and Campylobacter spp. to tolerate osmotic and desiccation stresses...... and identifies gaps in knowledge which need to be addressed to ensure the safety of low water activity and desiccated food products....

  17. Bioterrorism: toxins as weapons.

    Science.gov (United States)

    Anderson, Peter D

    2012-04-01

    The potential for biological weapons to be used in terrorism is a real possibility. Biological weapons include infectious agents and toxins. Toxins are poisons produced by living organisms. Toxins relevant to bioterrorism include ricin, botulinum, Clostridium perfrigens epsilson toxin, conotoxins, shigatoxins, saxitoxins, tetrodotoxins, mycotoxins, and nicotine. Toxins have properties of biological and chemical weapons. Unlike pathogens, toxins do not produce an infection. Ricin causes multiorgan toxicity by blocking protein synthesis. Botulinum blocks acetylcholine in the peripheral nervous system leading to muscle paralysis. Epsilon toxin damages cell membranes. Conotoxins block potassium and sodium channels in neurons. Shigatoxins inhibit protein synthesis and induce apoptosis. Saxitoxin and tetrodotoxin inhibit sodium channels in neurons. Mycotoxins include aflatoxins and trichothecenes. Aflatoxins are carcinogens. Trichothecenes inhibit protein and nucleic acid synthesis. Nicotine produces numerous nicotinic effects in the nervous system.

  18. RSV Test

    Science.gov (United States)

    ... Sex Hormone Binding Globulin (SHBG) Shiga toxin-producing Escherichia coli Sickle Cell Tests Sirolimus Smooth Muscle Antibody (SMA) ... University Medical Center, LAB Letter [On-line newsletter]. PDF available for download at http://www.stanfordhospital.com/ ...

  19. Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins

    OpenAIRE

    Mantzouki, Evanthia; Lürling, Miquel; Fastner, Jutta; de Senerpont Domis, Lisette; Wilk-Woźniak, Elżbieta; Koreivienė, Judita; Seelen, Laura; Teurlincx, Sven; Verstijnen, Yvon; Krztoń, Wojciech; Walusiak, Edward; Karosienė, Jūratė; Kasperovičienė, Jūratė; Savadova, Ksenija; Vitonytė, Irma

    2018-01-01

    Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins). Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a) and cytotoxins (e.g., cylindrospermopsin) due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and li...

  20. Temperature effects explain continental scale distribution of cyanobacterial toxins

    OpenAIRE

    Mantzouki, Evanthia; Lürling, Miquel; Fastner, Jutta; de Senerpont Domis, Lisette; Wilk-Woźniak, Elżbieta; Koreivienė, Judita; Seelen, Laura; Teurlincx, Sven; Verstijnen, Yvon; Krztoń, Wojciech; Walusiak, Edward; Karosienė, Jūratė; Kasperovičienė, Jūratė; Savadova, Ksenija; Vitonytė, Irma

    2018-01-01

    Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins). Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a) and cytotoxins (e.g., cylindrospermopsin) due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and li...

  1. Diarrhetic shellfish poisoning toxin esters in Danish blue mussels and surf clams

    DEFF Research Database (Denmark)

    Jørgensen, Kevin; Scanlon, Sine Hedegaard; Jensen, L.B.

    2005-01-01

    Until recently, little focus was given to the presence of diarrhetic shellfish poisoning ( DSP) toxin esters in seafood products. However, during the last few years, the occurrence of a high percentage of esters of the total amount of DSP toxins present in some seafood products has been observed....... importance because of the increased use of chemical methods instead of mouse bioassay for the detection of DSP toxicity....

  2. Prevalence and Characterization of a Binary Toxin (Actin-Specific ADP-Ribosyltransferase) from Clostridium difficile

    Science.gov (United States)

    Gonçalves, Carina; Decré, Dominique; Barbut, Frédéric; Burghoffer, Béatrice; Petit, Jean-Claude

    2004-01-01

    In addition to the two large clostridial cytotoxins (TcdA and TcdB), some strains of Clostridium difficile also produce an actin-specific ADP-ribosyltransferase, called binary toxin CDT. We used a PCR method and Southern blotting for the detection of genes encoding the enzymatic (CDTa) and binding (CDTb) components of the binary toxin in 369 strains isolated from patients with suspected C. difficile-associated diarrhea or colitis. Twenty-two strains (a prevalence of 6%) harbored both genes. When binary toxin production was assessed by Western blotting, 19 of the 22 strains reacted with antisera against the iota toxin of C. perfringens (anti-Ia and anti-Ib). Additionally, binary toxin activity, detected by the ADP-ribosyltransferase assay, was present in only 17 of the 22 strains. Subsequently, all 22 binary toxin-positive strains were tested for the production of toxins TcdA and TcdB, toxinotyped, and characterized by serogrouping, PCR ribotyping, arbitrarily primed PCR, and pulsed-field gel electrophoresis. All binary toxin-positive strains also produced TcdB and/or TcdA. However, they had significant changes in the tcdA and tcdB genes and belonged to variant toxinotypes III, IV, V, VII, IX, and XIII. We could differentiate 16 profiles by using typing methods, indicating that most of the binary toxin-positive strains were unrelated. PMID:15131151

  3. Phenol-Soluble Modulin Toxins of Staphylococcus haemolyticus

    Directory of Open Access Journals (Sweden)

    Fei Da

    2017-05-01

    Full Text Available Coagulase-negative staphylococci (CoNS are important nosocomial pathogens and the leading cause of sepsis. The second most frequently implicated species, after Staphylococcus epidermidis, is Staphylococcus haemolyticus. However, we have a significant lack of knowledge about what causes virulence of S. haemolyticus, as virulence factors of this pathogen have remained virtually unexplored. In contrast to the aggressive pathogen Staphylococcus aureus, toxin production has traditionally not been associated with CoNS. Recent findings have suggested that phenol-soluble modulins (PSMs, amphipathic peptide toxins with broad cytolytic activity, are widespread in staphylococci, but there has been no systematic assessment of PSM production in CoNS other than S. epidermidis. Here, we identified, purified, and characterized PSMs of S. haemolyticus. We found three PSMs of the β-type, which correspond to peptides that before were described to have anti-gonococcal activity. We also detected an α-type PSM that has not previously been described. Furthermore, we confirmed that S. haemolyticus does not produce a δ-toxin, as results from genome sequencing had indicated. All four S. haemolyticus PSMs had strong pro-inflammatory activity, promoting neutrophil chemotaxis. Notably, we identified in particular the novel α-type PSM, S. haemolyticus PSMα, as a potent hemolysin and leukocidin. For the first time, our study describes toxins of this important staphylococcal pathogen with the potential to have a significant impact on virulence during blood infection and sepsis.

  4. Influence of temperature shifts on survival, growth, and toxin production by psychrotrophic and mesophilic strains of Bacillus cereus in potatoes and chicken gravy.

    Science.gov (United States)

    Mahakarnchanakul, W; Beuchat, L R

    1999-03-15

    A study was done to determine the influence of temperature on growth and toxin production characteristics of psychrotrophic and mesophilic strains of Bacillus cereus when inoculated into mashed potatoes and chicken gravy containing various concentrations of sodium chloride and held at temperatures different from those at which cells had been cultured. Logarithmic growth phase cells (10 h, 30 degrees C) of psychrotrophic (F3802A/84) and mesophilic (B4ac-1) strains of Bacillus cereus were inoculated into rehydrated commercially processed instant mashed potatoes and chicken gravy supplemented with 0, 2, or 4% sodium chloride. Growth, survival, and diarrheal toxin production in potatoes and gravy held at 30, 37, and 10 degrees C (strain F3802A/84) or 30, 40, and 10 degrees C (strain B4ac-1) were monitored. Both strains grew in both foods containing no added sodium chloride or 2% sodium chloride when held at 30, 37, or 40 degrees C for 2 days. Strain B4ac-1 grew better than strain F3802A/84 in foods containing 4% sodium chloride. Maximum amounts of enterotoxin (1024 ng/g) were produced by strain B4ac-1 in chicken gravy held at 30 and 40 degrees C. Strain F3802A/84 grew to populations of 7 log10 CFU/g in foods containing no added sodium chloride or 2% sodium chloride at 10 degrees C. Strain F3802A/84 produced the highest amount of enterotoxin (1024 ng/g) at 30 degrees C in chicken gravy containing 0.7 or 2% sodium chloride; however, little or low amounts of toxin (4-16 ng/g) were produced in chicken gravy at 10 degrees C. Compared to strain B4ac-1, cells of strain F3802A/84 subjected to a downward shift in incubation temperature (10 degrees C) grew more rapidly in chicken gravy. Strain B4ac-1 produced the highest amount of toxin (1024 ng/g) at 30 degrees C in gravy containing 4% sodium chloride and at 40 degrees C in gravy containing 0.7% sodium chloride. Toxin was not detected in inoculated mashed potatoes. Results of this study indicate that shifts in incubation

  5. Studies on growth and toxin production of C. botulinum type E on cod homogenate treated with a combination of spices, sodium chloride and gamma-radiation

    International Nuclear Information System (INIS)

    Siddiqui, A.K.; Ando, Y.; Karashimada, T.; Kameyama, K.

    1979-01-01

    Cod homogenates inoculated with spores of C. botulinum type E strain Erimo at 10 2 and 10 4 /g were treated with 1% and 2% sodium chloride, 0.25% each of mustard, garlic and turmeric and 0.3 Mrad ν-radiation either in single or combination treatments. The growth and toxin production of type E spores in the inoculated homogenates were followed at incubation temperatures of 30 0 , 10 0 and 5 0 C for 7, 28 and 56 days respectively. Growth and gas formation were noted in all the samples but type E toxin could not be detected. The reason for the absence of toxin in both the untreated and treated homogenates could not be ascertained. Inadequate detection method, unfavourable growth conditions in the homogenate and weak toxigenicity of the strain employed have been advanced as probable factors that contributed to the negative results on the toxin assay. (author)

  6. Marine and freshwater toxins.

    Science.gov (United States)

    Hungerford, James M

    2006-01-01

    . (Abstracts from the Baiona 2005 meeting cited in this report can be found in the online version of the conference abstract book in the Files and Folders section of the Marine and Freshwater Toxins online community at www.aoac.org.) An active topic for discussion in Baiona and subsequent Task Force activities was the expert consultation for Codex which met in Oslo, Norway in 2004 (previously described and cited in last year's GR report, ref 1). The consultation group's executive summary report (http://www.fao.org/es/ESN/food/risk_biotoxin en.stm) describes suggested changes in action levels as well as methods, method validation, and other issues. September 2005 saw the AOAC Task Force efforts further supported by another symposium, "Marine and Freshwater Toxins: Quality Methods for Food Safety and International Trade," at the AOAC INTERNATIONAL Annual Conference in Orlando, Florida. The multidisciplinary talks at this full day symposium ranged from ciguatoxins to cyanobacterial toxins, and spanned toxicology, biochemistry, molecular biology and analytical chemistry. Again, the symposium preceded Task Force meetings. Toxin subgroups, including a new group on cyanobacterial toxins, met for engaging and productive subgroup discussions. All of these activities were preceded by a Wiley Award symposium for Task Force member Mike Quilliam of NRC Canada. These talks, presented at a half-day symposium on the first day of the Annual Meeting, focused on Quilliam's work with LC tandem mass spectrometry (LC/MS/MS) and certified reference standards and materials, and included related presentations by some of his many research collaborators. To maintain flow and continuity between symposia and between Task Force meetings, the group now uses new electronic discussion forums. Individual subgroup areas, under the Marine and Freshwater Toxins Task Force, comprise this online community. First introduced by AOAC INTERNATIONAL in early 2005, these new resources are being used to distribute

  7. Differentiation and distribution of three types of exfoliative toxin produced by Staphylococcus hyicus from pigs with exudative epidermitis

    DEFF Research Database (Denmark)

    Andresen, Lars Ole

    1998-01-01

    were antigenically distinct. The three toxins were designated ExhA, ExhB and ExhC. From 60 diseased pigs, each representing an outbreak of exudative epidermitis, a total of 584 isolates of S. hl icus were phage typed and tested for production of exfoliative toxin. ExhA-, ExhB- and ExhC-producing S....... hyicus isolates were found in 12 (20%), 20 (33%) and 11 (18%); respectively, of the 60 pig herds investigated. Production of the different types of exfoliative toxin was predominantly associated with certain phage groups. However. toxin production was found in all of the six phage groups defined...

  8. Botulinum toxin

    Directory of Open Access Journals (Sweden)

    Nigam P

    2010-01-01

    Full Text Available Botulinum toxin, one of the most poisonous biological substances known, is a neurotoxin produced by the bacterium Clostridium botulinum. C. botulinum elaborates eight antigenically distinguishable exotoxins (A, B, C 1 , C 2 , D, E, F and G. All serotypes interfere with neural transmission by blocking the release of acetylcholine, the principal neurotransmitter at the neuromuscular junction, causing muscle paralysis. The weakness induced by injection with botulinum toxin A usually lasts about three months. Botulinum toxins now play a very significant role in the management of a wide variety of medical conditions, especially strabismus and focal dystonias, hemifacial spasm, and various spastic movement disorders, headaches, hypersalivation, hyperhidrosis, and some chronic conditions that respond only partially to medical treatment. The list of possible new indications is rapidly expanding. The cosmetological applications include correction of lines, creases and wrinkling all over the face, chin, neck, and chest to dermatological applications such as hyperhidrosis. Injections with botulinum toxin are generally well tolerated and side effects are few. A precise knowledge and understanding of the functional anatomy of the mimetic muscles is absolutely necessary to correctly use botulinum toxins in clinical practice.

  9. [Active etiological surveillance for foodborne diseases in Guangdong province, 2013-2014

    NARCIS (Netherlands)

    Ke, B. X.; He, D. M.; Tan, H. L.; Zeng, H. H.; Yang, T.; Li, B. S.; Liang, Y. H.; Lu, L. L.; Liang, J. H.; Huang, Q.; Ke, C. W.

    2016-01-01

    Objective: To study the infection status, serotypes, drug resistance and molecular characteristics of Salmonella, Shigella, Vibrio parahemolyticus, enterotoxigenic Escherichia (E.) coli (ETEC), pathogenic E. coli (EPEC), Shiga Toxin producing E. coli (STEC) and Enteroinvasive E. coli (EIEC)

  10. Factors affecting growth and toxin production by Clostridium botulinum type E on irradiated (0.3 Mrad) chicken skins

    International Nuclear Information System (INIS)

    Firstenberg-Eden, R.; Rowley, D.B.; Shattuck, G.E.

    1982-01-01

    A model system (chicken skins with chicken exudate) was used to determine if Clostridium botulinum type E (Beluga) spores, stressed by low dose irradiation, would develop and produce toxin at abuse temperatures of 10 and 30 0 C in the absence of characteristic spoilage. Unstressed spores germinated, multiplied, and produced toxin on vacuum-packed chicken skins, stored at either 30 or 10 0 C. Cell numbers increased faster and toxin was evident sooner at 30 0 C than at 10 0 C. At 30 0 C, growth occurred and toxin was produced more slowly when samples were incubated aerobically than anaerobically. When samples were incubated aerobically at 10 0 C, no toxin was detected within a test period of 14 days. An irradiation dose of 0.3 Mrad at 5 0 C reduced a spore population on vacuum-sealed chicken skins by about 90%. The surviving population produced toxin at 30 0 C under either aerobic or anaerobic conditions, at 10 0 C no toxin was detected even on skins incubated anaerobically. Under the worst conditions (30 0 C, vacuum packed) toxin was not detected prior to characteristic spoilage caused by the natural flora surviving 0.3 Mrad

  11. A thiol-disulfide oxidoreductase of the Gram-positive pathogen Corynebacterium diphtheriae is essential for viability, pilus assembly, toxin production and virulence.

    Science.gov (United States)

    Reardon-Robinson, Melissa E; Osipiuk, Jerzy; Jooya, Neda; Chang, Chungyu; Joachimiak, Andrzej; Das, Asis; Ton-That, Hung

    2015-12-01

    The Gram-positive pathogen Corynebacterium diphtheriae exports through the Sec apparatus many extracellular proteins that include the key virulence factors diphtheria toxin and the adhesive pili. How these proteins attain their native conformations after translocation as unfolded precursors remains elusive. The fact that the majority of these exported proteins contain multiple cysteine residues and that several membrane-bound oxidoreductases are encoded in the corynebacterial genome suggests the existence of an oxidative protein-folding pathway in this organism. Here we show that the shaft pilin SpaA harbors a disulfide bond in vivo and alanine substitution of these cysteines abrogates SpaA polymerization and leads to the secretion of degraded SpaA peptides. We then identified a thiol-disulfide oxidoreductase (MdbA), whose structure exhibits a conserved thioredoxin-like domain with a CPHC active site. Remarkably, deletion of mdbA results in a severe temperature-sensitive cell division phenotype. This mutant also fails to assemble pilus structures and is greatly defective in toxin production. Consistent with these defects, the ΔmdbA mutant is attenuated in a guinea pig model of diphtheritic toxemia. Given its diverse cellular functions in cell division, pilus assembly and toxin production, we propose that MdbA is a component of the general oxidative folding machine in C. diphtheriae. © 2015 John Wiley & Sons Ltd.

  12. Studies on growth and toxin production of C. botulinum type E on cod homogenate treated with a combination of spices, sodium chloride and gamma-radiation

    Energy Technology Data Exchange (ETDEWEB)

    Siddiqui, A K [Atomic Energy Centre, Dacca (Bangladesh); Ando, Y; Karashimada, T; Kameyama, K

    1979-09-01

    Cod homogenates inoculated with spores of C. botulinum type E strain Erimo at 10/sup 2/ and 10/sup 4//g were treated with 1% and 2% sodium chloride, 0.25% each of mustard, garlic and turmeric and 0.3 Mrad ..gamma..-radiation either in single or combination treatments. The growth and toxin production of type E spores in the inoculated homogenates were followed at incubation temperatures of 30/sup 0/, 10/sup 0/ and 5/sup 0/C for 7, 28 and 56 days respectively. Growth and gas formation were noted in all the samples but type E toxin could not be detected. The reason for the absence of toxin in both the untreated and treated homogenates could not be ascertained. Inadequate detection method, unfavourable growth conditions in the homogenate and weak toxigenicity of the strain employed have been advanced as probable factors that contributed to the negative results on the toxin assay.

  13. Incidence of Escherichia coli O157:H7 in Thailand

    International Nuclear Information System (INIS)

    Sukhumungoon, P.

    2015-01-01

    Entero hemorrhagic Escherichia coli (EHEC) especially serotype O157:H7 is one of the important food-borne pathogens because it is able to produce crucial toxins Shiga. However, the outbreak of this organism in Thailand has not been reported. Antibody to O157 antigen was detected in some Thai populations and Shiga toxin-producing E. coli were detected in low numbers of clinical specimens. Interestingly, some E. coli that showed positive to O157 fimbriae probe and lack of virulence gene were isolated from certain patients and one isolate of E. coli O157:H7 which possessed stx1, stx2v was detected in a normal child. In addition, the incidence of E. coli O157:H7 strains were monitored by the samples from cattle and retail beef in Thailand although their inability to produce toxins or produce in a low concentration was demonstrated. This review discusses the incidences of E. coli O157 in clinical and environmental samples of Thailand including the transmission possibility of this bacterium across the Thai border through food trade. (author)

  14. Characterization of the SigD regulon of C. difficile and its positive control of toxin production through the regulation of tcdR.

    Directory of Open Access Journals (Sweden)

    Imane El Meouche

    Full Text Available Clostridium difficile intestinal disease is mediated largely by the actions of toxins A (TcdA and B (TcdB, whose production occurs after the initial steps of colonization involving different surface or flagellar proteins. In B. subtilis, the sigma factor SigD controls flagellar synthesis, motility, and vegetative autolysins. A homolog of SigD encoding gene is present in the C.difficile 630 genome. We constructed a sigD mutant in C. difficile 630 ∆erm to analyze the regulon of SigD using a global transcriptomic approach. A total of 103 genes were differentially expressed between the wild-type and the sigD mutant, including genes involved in motility, metabolism and regulation. In addition, the sigD mutant displayed decreased expression of genes involved in flagellar biosynthesis, and also of genes encoding TcdA and TcdB as well as TcdR, the positive regulator of the toxins. Genomic analysis and RACE-PCR experiments allowed us to characterize promoter sequences of direct target genes of SigD including tcdR and to identify the SigD consensus. We then established that SigD positively regulates toxin expression via direct control of tcdR transcription. Interestingly, the overexpression of FlgM, a putative anti-SigD factor, inhibited the positive regulation of motility and toxin synthesis by SigD. Thus, SigD appears to be the first positive regulator of the toxin synthesis in C. difficile.

  15. Botulinum toxin in parkinsonism: The when, how, and which for botulinum toxin injections.

    Science.gov (United States)

    Cardoso, Francisco

    2018-06-01

    The aim of this article is to provide a review of the use of injections of botulinum toxin in the management of selected symptoms and signs of Parkinson's disease and other forms of parkinsonism. Sialorrhea is defined as inability to control oral secretions, resulting in excessive saliva in the oropharynx. There is a high level of evidence for the treatment of sialorrhea in parkinsonism with injections of different forms of botulinum toxin type A as well as botulinum toxin type B. Tremor can be improved by the use of botulinum toxin injections but improved tremor control often leads to concomitant motor weakness, limiting its use. Levodopa induced dyskinesias are difficult to treat with botulinum toxin injections because of their variable frequency and direction. Apraxia of eyelid opening, a sign more commonly seen in progressive supranuclear palsy and other tauopathies, often improves after botulinum toxin injections. Recent data suggest that regardless of the underlying mechanism, pain in parkinsonism can be alleviated by botulinum toxin injections. Finally, freezing of gait, camptocormia and Pisa syndrome in parkinsonism almost invariably fail to respond to botulinum toxin injections. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. The effects of globotriaosylceramide tail saturation level on bilayer phases

    DEFF Research Database (Denmark)

    Pezeshkian, Weria; Chaban, Vitaly V; Johannes, Ludger

    2015-01-01

    Globotriaosylceramide (Gb3) is a glycosphingolipid present in the plasma membrane that is the natural receptor of the bacterial Shiga toxin. The unsaturation level of Gb3 acyl chains has a drastic impact on lipid bilayer properties and phase behaviour, and on many Gb3-related cellular processes....... For example: the Shiga toxin B subunit forms tubular invaginations in the presence of Gb3 with an unsaturated acyl chain (U-Gb3), while in the presence of Gb3 with a saturated acyl chain (S-Gb3) such invagination does not occur. We have used all-atom molecular dynamics simulations to investigate the effects...... of sphingomyelin lipids and (3) At higher Gb3 concentrations, U-Gb3 mixes better with dioleoylphosphatidylcholine than S-Gb3. Our simulations also provide the first molecular level structural model of Gb3 in membranes....

  17. Growth of and toxin production by nonproteolytic Clostridium botulinum in cooked puréed vegetables at refrigeration temperatures.

    Science.gov (United States)

    Carlin, F; Peck, M W

    1996-01-01

    Seven strains of nonproteolytic Clostridium botulinum (types B, E, and F) were each inoculated into a range of anaerobic cooked puréed vegetables. After incubation at 10 degrees C for 15 to 60 days, all seven strains formed toxin in mushrooms, five did so in broccoli, four did so in cauliflower, three did so in asparagus, and one did so in kale. Growth kinetics of nonproteolytic C. botulinum type B in cooked mushrooms, cauliflower, and potatoes were determined at 16, 10, 8, and 5 degrees C. Growth and toxin production occurred in cooked cauliflower and mushrooms at all temperatures and in potatoes at 16 and 8 degrees C. The C. botulinum neurotoxin was detected within 3 to 5 days at 16 degrees C, 11 to 13 days at 10 degrees C, 10 to 34 days at 8 degrees C, and 17 to 20 days at 5 degrees C. PMID:8702303

  18. Fumonisin and T-2 toxin production of Fusarium spp. isolated from complete feed and individual agricultural commodities used in shrimp farming.

    Science.gov (United States)

    Anukul, Nampeung; Maneeboon, Thanapoom; Roopkham, Chanram; Chuaysrinule, Chananya; Mahakarnchanakul, Warapa

    2014-02-01

    Fusarium spp. are plant pathogens producing fumonisins and trichothecenes that both affect human and animal health. In the present study, 40 fungal strains were isolated and species identified from 35 shrimp feed samples and from 61 agricultural raw materials. F. verticillioides was the predominant species (85 %) mostly found in corn and soybean meal, while no Fusarium contamination was detected in shrimp feed. Levels of 10 % of F. oxysporum were isolated from peanut and 5 % of F. equiseti contamination in corn and peanut. To determine the ability of toxin production, enzyme-linked immunosorbent assay, polymerase chain reaction, and ultra-pressure liquid chromatography-tandem mass spectrometry were performed. All but four of the fumonisin-producing strains contained the FUM1 gene. No Fusarium synthesized T-2 toxin nor contained the Tri5 gene. This survey brings more data on mycotoxin contamination in the food chain of animal feed production, and leads to the awareness of the use of contaminated raw materials in shrimp farming.

  19. Eeyarestatin 1 interferes with both retrograde and anterograde intracellular trafficking pathways.

    Directory of Open Access Journals (Sweden)

    Mina-Olga Aletrari

    Full Text Available The small molecule Eeyarestatin I (ESI inhibits the endoplasmic reticulum (ER-cytosol dislocation and subsequent degradation of ERAD (ER associated protein degradation substrates. Toxins such as ricin and Shiga/Shiga-like toxins (SLTx are endocytosed and trafficked to the ER. Their catalytic subunits are thought to utilise ERAD-like mechanisms to dislocate from the ER into the cytosol, where a proportion uncouples from the ERAD process, recovers a catalytic conformation and destroys their cellular targets. We therefore investigated ESI as a potential inhibitor of toxin dislocation.Using cytotoxicity measurements, we found no role for ES(I as an inhibitor of toxin dislocation from the ER, but instead found that for SLTx, ESI treatment of cells was protective by reducing the rate of toxin delivery to the ER. Microscopy of the trafficking of labelled SLTx and its B chain (lacking the toxic A chain showed a delay in its accumulation at a peri-nuclear location, confirmed to be the Golgi by examination of SLTx B chain metabolically labelled in the trans-Golgi cisternae. The drug also reduced the rate of endosomal trafficking of diphtheria toxin, which enters the cytosol from acidified endosomes, and delayed the Golgi-specific glycan modifications and eventual plasma membrane appearance of tsO45 VSV-G protein, a classical marker for anterograde trafficking.ESI acts on one or more components that function during vesicular transport, whilst at least one retrograde trafficking pathway, that of ricin, remains unperturbed.

  20. Polyamine toxins

    DEFF Research Database (Denmark)

    Strømgaard, Kristian; Jensen, Lars S; Vogensen, Stine B

    2005-01-01

    Polyamine toxins, isolated from spiders and wasps, have been used as pharmacological tools for the study of ionotropic receptors, but their use have so far been hampered by their lack of selectivity. In this mini-review, we describe how careful synthetic modification of native polyamine toxins ha...

  1. Botulinum toxin type A products are not interchangeable: a review of the evidence

    Directory of Open Access Journals (Sweden)

    Brin MF

    2014-10-01

    Full Text Available Mitchell F Brin,1,2 Charmaine James,3 John Maltman1 1Allergan, Inc., Irvine, CA, USA; 2Department of Neurology, University of California, Irvine, CA, USA; 3Allergan, Marlow, UKAbstract: Botulinum toxin type A (BoNTA products are injectable biologic medications derived from Clostridium botulinum bacteria. Several different BoNTA products are marketed in various countries, and they are not interchangeable. Differences between products include manufacturing processes, formulations, and the assay methods used to determine units of biological activity. These differences result in a specific set of interactions between each BoNTA product and the tissue injected. Consequently, the products show differences in their in vivo profiles, including preclinical dose response curves and clinical dosing, efficacy, duration, and safety/adverse events. Most, but not all, published studies document these differences, suggesting that individual BoNTA products act differently depending on experimental and clinical conditions, and these differences may not always be predictable. Differentiation through regulatory approvals provides a measure of confidence in safety and efficacy at the specified doses for each approved indication. Moreover, the products differ in the amount of study to which they have been subjected, as evidenced by the number of publications in the peer-reviewed literature and the quantity and quality of clinical studies. Given that BoNTAs are potent biological products that meet important clinical needs, it is critical to recognize that their dosing and product performance are not interchangeable and each product should be used according to manufacturer guidelines.Keywords: onabotulinumtoxinA, abobotulinumtoxinA, incobotulinumtoxinA, glabellar lines, non-interchangeability

  2. Identification of serotypes and virulence markers of Escherichia coli isolated from human stool and urine samples in Egypt

    Directory of Open Access Journals (Sweden)

    K M Osman

    2012-01-01

    Full Text Available Purpose: Haemorrhagic colitis and haemolytic-uremic syndrome are associated with Shiga-toxin producing Escherichia coli (STEC. There are others DEC (Diarrhoeagenic E. coli pathotypes responsible for outbreaks and others toxins associated to these. Most clinical signs of disease arise as a consequence of the production of Shiga toxin 1 (Stx1, Stx2 or combinations of these toxins. Other major virulence factors include E. coli haemolysin (hlyA, and intimin, the product of the eaeA gene that is involved in the attaching and effacing adherence phenotype. Materials and Methods: In this study, the PCR assay was used to detect 12 E. coli genes associated with virulence (stx1, stx2, hylA, Flic h7 , stb, F41, K99, sta, F17, LT-I, LT-II and eaeA. Results: A total of 108 E. coli strains were serotyped into 64 typable strains. The investigated strains from the stool, 8/80 (10% strains were O 164:K, while the 56/110 strains isolated from the urine were O126:K71 (44/110, 40% and O 86:K 61 (12/110, 11%. The distribution pattern of the detected virulence genes was observed to be in the following order: F17 (10% from the stool and 44% from the urine, Sta (10% from the stool, hylA (10% from the stool and 44% from the urine, Stb (44% from the urine and stx1 (27% from the urine. The 8 faecal strains encoded a combination of the F17, Sta and hylA genes, while the 56 urine strains encoded a combination of the F17 0+ Stb + hylA (44/110, 40% and Stx1 only (12/60, 20%. Conclusion: This is the first report on the molecular characterization of E. coli diarrhoeagenic strains in Egypt and the first report on the potential role of E. coli in diarrhoea and urinary tract infections in a localized geographic area where the people engage in various occupational activities.

  3. Identification and apoptotic potential of T-2 toxin metabolites in human cells

    NARCIS (Netherlands)

    Weidner, M.; Welsch, T.; Hübner, F.; Schwerdt, G.; Gekle, M.; Humpf, H.U.

    2012-01-01

    The mycotoxin T-2 toxin, produced by various Fusarium species, is a widespread contaminant of grain and grain products. Knowledge about its toxicity and metabolism in the human body is crucial for any risk assessment as T-2 toxin can be detected in processed and unprocessed food samples. Cell

  4. Higher cytotoxicity of divalent antibody-toxins than monovalent antibody-toxins

    International Nuclear Information System (INIS)

    Won, JaeSeon; Nam, PilWon; Lee, YongChan; Choe, MuHyeon

    2009-01-01

    Recombinant antibody-toxins are constructed via the fusion of a 'carcinoma-specific' antibody fragment to a toxin. Due to the high affinity and high selectivity of the antibody fragments, antibody-toxins can bind to surface antigens on cancer cells and kill them without harming normal cells [L.H. Pai, J.K. Batra, D.J. FitzGerald, M.C. Willingham, I. Pastan, Anti-tumor activities of immunotoxins made of monoclonal antibody B3 and various forms of Pseudomonas exotoxin, Proc. Natl. Acad. Sci. USA 88 (1991) 3358-3362]. In this study, we constructed the antibody-toxin, Fab-SWn-PE38, with SWn (n = 3, 6, 9) sequences containing n-time repeated (G 4 S) between the Fab fragment and PE38 (38 kDa truncated form of Pseudomonas exotoxin A). The SWn sequence also harbored one cysteine residue that could form a disulfide bridge between two Fab-SWn-PE38 monomers. We assessed the cytotoxicity of the monovalent (Fab-SWn-PE38), and divalent ([Fab-SWn-PE38] 2 ) antibody-toxins. The cytotoxicity of the dimer against the CRL1739 cell line was approximately 18.8-fold higher than that of the monomer on the ng/ml scale, which was approximately 37.6-fold higher on the pM scale. These results strongly indicate that divalency provides higher cytotoxicity for an antibody-toxin.

  5. Detección, aislamiento y caracterización de Escherichia coli productor de toxina Shiga a partir de carne molida fresca proveniente de carnicerías de Concepción, provincia de Tucumán Detection, isolation and characterization of Shiga toxin-producing Escherichia coli (STEC in fresh ground beef from butcher shops in Concepción, Tucumán Province

    Directory of Open Access Journals (Sweden)

    M. A. Jure

    2010-12-01

    Full Text Available Escherichia coli productor de toxina Shiga (STEC es un patógeno emergente transmitido por alimentos. Existen numerosos serotipos de STEC asociados a enfermedad en humanos, entre los cuales prevalece el serotipo O157:H7. La carne molida es el principal vehículo de transmisión. En la ciudad de Concepción, provincia de Tucumán, entre setiembre y diciembre de 2004 se diagnosticaron dos casos de síndrome urémico hemolítico (SUH. El objetivo de este trabajo fue detectar, aislar y caracterizar STEC O157 y no-O157 a partir de muestras de carne molida fresca obtenidas en las bocas de expendio. Entre los meses de setiembre y diciembre de 2004 se recolectaron 53 muestras de carne molida fresca en carnicerías de la ciudad de Concepción. Para la detección, el aislamiento y la caracterización de STEC O157:H7 se utilizó la metodología USDA-FSIS 2002. Para la detección de E. coli no-O157 se utilizaron dos técnicas de PCR; para el aislamiento y la caracterización se utilizó una metodología previamente validada en una etapa intralaboratorio. Siete muestras fueron positivas para el gen stx2, de las cuales 4 también fueron positivas para el gen rfbO157. Sin embargo, solo se aisló una cepa de E. coli O157:H7 biotipo C, portadora de los genes eae, stx2 y ehxA. El presente trabajo refleja la importancia de implementar técnicas que permitan detectar este grupo de patógenos emergentes a partir de productos cárnicos.Shiga toxin-producing Escherichia coli is an emerging foodborne pathogen. There are many STEC serotypes associated with human diseases, being the O157:H7 serotype the most prevalent. Ground beef is the main transmission vehicle. In Concepción city, Tucumán Province, between September and December 2004, two hemolytic uremic syndrome (HUS cases were diagnosed. The main objective of this work was to detect, isolate and characterize STEC O157 and non-O157 strains in fresh ground beef. Between September and December 2004, 53 fresh ground

  6. Complement Test

    Science.gov (United States)

    ... Salicylates Semen Analysis Serotonin Serum Free Light Chains Sex Hormone Binding Globulin (SHBG) Shiga toxin-producing Escherichia ... and forming complexes that respond to infections, non-self tissues (transplants), dead cells ... KJ. Complement determinations in human disease. Ann Allergy Asthma Immunol . 2004; ...

  7. Shigella-specific IgA in saliva of children with bacillary dysentery

    NARCIS (Netherlands)

    Schultsz, C.; Qadri, F.; Hossain, S. A.; Ahmed, F.; Ciznar, I.

    1992-01-01

    To study the secretory immune response after Shigella infection, the anti-lipopolysaccharide and anti-Shiga-toxin response in saliva, obtained from children with confirmed shigellosis and healthy children, were determined by enzyme-linked immunosorbent assay and by Western blot. Children with

  8. Monoclonal Antibodies and Toxins—A Perspective on Function and Isotype

    Directory of Open Access Journals (Sweden)

    Siu-Kei Chow

    2012-06-01

    Full Text Available Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins—Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB—and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions.

  9. Monoclonal Antibodies and Toxins—A Perspective on Function and Isotype

    Science.gov (United States)

    Chow, Siu-Kei; Casadevall, Arturo

    2012-01-01

    Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins—Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)—and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions. PMID:22822456

  10. Autoproteolytic Activation of Bacterial Toxins

    Directory of Open Access Journals (Sweden)

    Aimee Shen

    2010-05-01

    Full Text Available Protease domains within toxins typically act as the primary effector domain within target cells. By contrast, the primary function of the cysteine protease domain (CPD in Multifunctional Autoprocessing RTX-like (MARTX and Clostridium sp. glucosylating toxin families is to proteolytically cleave the toxin and release its cognate effector domains. The CPD becomes activated upon binding to the eukaryotic-specific small molecule, inositol hexakisphosphate (InsP6, which is found abundantly in the eukaryotic cytosol. This property allows the CPD to spatially and temporally regulate toxin activation, making it a prime candidate for developing anti-toxin therapeutics. In this review, we summarize recent findings related to defining the regulation of toxin function by the CPD and the development of inhibitors to prevent CPD-mediated activation of bacterial toxins.

  11. Conjugated Linoleic Acid Reduces Cholera Toxin Production In Vitro and In Vivo by Inhibiting Vibrio cholerae ToxT Activity.

    Science.gov (United States)

    Withey, Jeffrey H; Nag, Drubhajyoti; Plecha, Sarah C; Sinha, Ritam; Koley, Hemanta

    2015-12-01

    The severe diarrheal disease cholera is endemic in over 50 countries. Current therapies for cholera patients involve oral and/or intravenous rehydration, often combined with the use of antibiotics to shorten the duration and intensity of the disease. However, as antibiotic resistance increases, treatment options will become limited. Linoleic acid has been shown to be a potent negative effector of V. cholerae virulence that acts on the major virulence transcription regulator protein, ToxT, to inhibit virulence gene expression. ToxT activates transcription of the two major virulence factors required for disease, cholera toxin (CT) and toxin-coregulated pilus (TCP). A conjugated form of linoleic acid (CLA) is currently sold over the counter as a dietary supplement and is generally recognized as safe by the U.S. Food and Drug Administration. This study examined whether CLA could be used as a new therapy to reduce CT production, which, in turn, would decrease disease duration and intensity in cholera patients. CLA could be used in place of traditional antibiotics and would be very unlikely to generate resistance, as it affects only virulence factor production and not bacterial growth or survival. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. Dietary toxins, endoplasmic reticulum (ER) stress and diabetes.

    Science.gov (United States)

    Hettiarachchi, Kalindi D; Zimmet, Paul Z; Myers, Mark A

    2008-05-01

    The incidence of Type 1 diabetes has been increasing at a rate too rapid to be due to changes in genetic risk. Instead changes in environmental factors are the likely culprit. The endoplasmic reticulum (ER) plays an important role in the production of newly synthesized proteins and interference with these processes leads to ER stress. The insulin-producing beta cells are particularly prone to ER stress as a result of their heavy engagement in insulin production. Increasing evidence suggests ER stress is central to initiation and progression of Type 1 diabetes. An early environmental exposure, such as toxins and viral infections, can impart a significant physiological load on beta cells to initiate abnormal processing of proinsulin, ER stress and insulin secretory defects. Release of altered proinsulin from the beta cells early in life may trigger autoimmunity in those with genetic susceptibility leading to cytokine-induced nitric oxide production and so exacerbating ER stress in beta cells, ultimately leading to apoptosis of beta cells and diabetes. Here we suggest that ER stress is an inherent cause of beta cell dysfunction and environmental factors, in particular dietary toxins derived from Streptomyces in infected root vegetables, can impart additional stress that aggravates beta cell death and progression to diabetes. Furthermore, we propose that the increasing incidence of Type 1 diabetes may be accounted for by increased dietary exposure to ER-stress-inducing Streptomyces toxins.

  13. A simple electroelution method for rapid protein purification: isolation and antibody production of alpha toxin from Clostridium septicum

    Directory of Open Access Journals (Sweden)

    Lorena Vázquez-Iglesias

    2017-06-01

    Full Text Available Clostridium septicum produces a number of diseases in human and farm animals which, in most of the cases, are fatal without clinical intervention. Alpha toxin is an important agent and the unique lethal virulent factor produced by Clostridium septicum. This toxin is haemolytic, highly lethal and necrotizing activities but is being used as an antigen to develop animal vaccines. The aim of this study was to isolate the alpha toxin of Clostridium septicum and produce highly specific antibodies against it. In this work, we have developed a simple and efficient method for alpha toxin purification, based on electroelution that can be used as a time-saving method for purifying proteins. This technique avoids contamination by other proteins that could appear during other protein purification techniques such chromatography. The highly purified toxin was used to produce polyclonal antibodies. The specificity of the antibodies was tested by western blot and these antibodies can be applied to the quantitative determination of alpha toxin by slot blot.

  14. A Kinetic and Factorial Approach to Study the Effects of Temperature and Salinity on Growth and Toxin Production by the Dinoflagellate Alexandrium ostenfeldii from the Baltic Sea.

    Directory of Open Access Journals (Sweden)

    Pablo Salgado

    Full Text Available Alexandrium ostenfeldii is present in a wide variety of environments in coastal areas worldwide and is the only dinoflagellate known species that produces paralytic shellfish poisoning (PSP toxins and two types of cyclic imines, spirolides (SPXs and gymnodimines (GYMs. The increasing frequency of A. ostenfeldii blooms in the Baltic Sea has been attributed to the warming water in this region. To learn more about the optimal environmental conditions favoring the proliferation of A. ostenfeldii and its complex toxicity, the effects of temperature and salinity on the kinetics of both the growth and the net toxin production of this species were examined using a factorial design and a response-surface analysis (RSA. The results showed that the growth of Baltic A. ostenfeldii occurs over a wide range of temperatures and salinities (12.5-25.5°C and 5-21, respectively, with optimal growth conditions achieved at a temperature of 25.5°C and a salinity of 11.2. Together with the finding that a salinity > 21 was the only growth-limiting factor detected for this strain, this study provides important insights into the autecology and population distribution of this species in the Baltic Sea. The presence of PSP toxins, including gonyautoxin (GTX-3, GTX-2, and saxitoxin (STX, and GYMs (GYM-A and GYM-B/-C analogues was detected under all temperature and salinity conditions tested and in the majority of the cases was concomitant with both the exponential growth and stationary phases of the dinoflagellate's growth cycle. Toxin concentrations were maximal at temperatures and salinities of 20.9°C and 17 for the GYM-A analogue and > 19°C and 15 for PSP toxins, respectively. The ecological implications of the optimal conditions for growth and toxin production of A. ostenfeldii in the Baltic Sea are discussed.

  15. 77 FR 31975 - Shiga Toxin-Producing Escherichia coli in Certain Raw Beef Products

    Science.gov (United States)

    2012-05-31

    ... International, the French Association for Standardization (AFNOR), the European organization for the validation... correlates with the concentration or frequency of another organism. FSIS-Recommended Cooking Temperatures Comment: One commenter stated that if STECs can survive ``ordinary'' or ``typical'' cooking, FSIS should...

  16. Bacteriophage-encoded shiga toxin gene in atypical bacterial host

    Directory of Open Access Journals (Sweden)

    Casas Veronica

    2011-07-01

    Full Text Available Abstract Background Contamination from fecal bacteria in recreational waters is a major health concern since bacteria capable of causing human disease can be found in animal feces. The Dog Beach area of Ocean Beach in San Diego, California is a beach prone to closures due to high levels of fecal indicator bacteria (FIB. A potential source of these FIB could be the canine feces left behind by owners who do not clean up after their pets. We tested this hypothesis by screening the DNA isolated from canine feces for the bacteriophage-encoded stx gene normally found in the virulent strains of the fecal bacterium Escherichia coli. Results Twenty canine fecal samples were collected, processed for total and bacterial fraction DNA, and screened by PCR for the stx gene. The stx gene was detected in the total and bacterial fraction DNA of one fecal sample. Bacterial isolates were then cultivated from the stx-positive fecal sample. Eighty nine of these canine fecal bacterial isolates were screened by PCR for the stx gene. The stx gene was detected in five of these isolates. Sequencing and phylogenetic analyses of 16S rRNA gene PCR products from the canine fecal bacterial isolates indicated that they were Enterococcus and not E. coli. Conclusions The bacteriophage-encoded stx gene was found in multiple species of bacteria cultivated from canine fecal samples gathered at the shoreline of the Dog Beach area of Ocean Beach in San Diego, California. The canine fecal bacteria carrying the stx gene were not the typical E. coli host and were instead identified through phylogenetic analyses as Enterococcus. This suggests a large degree of horizontal gene transfer of exotoxin genes in recreational waters.

  17. Toxin-Based Therapeutic Approaches

    Science.gov (United States)

    Shapira, Assaf; Benhar, Itai

    2010-01-01

    Protein toxins confer a defense against predation/grazing or a superior pathogenic competence upon the producing organism. Such toxins have been perfected through evolution in poisonous animals/plants and pathogenic bacteria. Over the past five decades, a lot of effort has been invested in studying their mechanism of action, the way they contribute to pathogenicity and in the development of antidotes that neutralize their action. In parallel, many research groups turned to explore the pharmaceutical potential of such toxins when they are used to efficiently impair essential cellular processes and/or damage the integrity of their target cells. The following review summarizes major advances in the field of toxin based therapeutics and offers a comprehensive description of the mode of action of each applied toxin. PMID:22069564

  18. Affinity chromatography of tetanus toxin, tetanus toxoid, and botulinum A toxin on synaptosomes, and differentiation of their acceptors

    Energy Technology Data Exchange (ETDEWEB)

    Habermann, E [Giessen Univ. (Germany, F.R.). Pharmakologisches Inst.

    1976-01-01

    /sup 125/I-labelled tetanus toxin and /sup 125/I-labelled botulinum A neurotoxin are known to be specifically bound to brain synaptosomes. In order to discriminate between active toxin and inactive admixtures present in the starting material or arising during iodination, synaptosome columns were prepared using bromacetylcellulose and/or kieselgur (Celite) as carriers. Both types of columns adsorb the toxins from low ionic strength medium and release them if the pH and ionic strength are raised. Botulinum toxin was eluted with lower ionic strength than tetanus toxin, and could be freed from nontoxic admixtures. Analysis by affinity chromatography disclosed partially toxoided tetanus toxin in both labelled and unlabelled toxin samples. High concentrations of formaldehyde (0.5%) destroyed both toxicity and affinity to the synaptosomes of tetanus toxin. Low concentrations of formaldehyde (0.05%) yielded a derivative of low toxicity which was still, however less firmly, bound to synaptosomes. Tetanus and botulinum toxin differ by their acceptors. Whereas unlabelled botulinum toxin is unable to compete with labelled tetanus toxin, unlabelled tetanus toxin slightly competes with botulinum toxin. Both labelled toxins display anomalous binding behaviour in that they cannot be displaced completely even with a large excess of unlabelled toxin.

  19. Affinity chromatography of tetanus toxin, tetanus toxoid, and botulinum A toxin on synaptosomes, and differentiation of their acceptors

    International Nuclear Information System (INIS)

    Habermann, E.

    1976-01-01

    125 I-labelled tetanus toxin and 125 I-labelled botulinum A neurotoxin are known to be specifically bound to brain synaptosomes. In order to discriminate between active toxin and inactive admixtures present in the starting material or arising during iodination, synaptosome columns were prepared using bromacetylcellulose and/or kieselgur (Celite) as carriers. Both types of columns adsorb the toxins from low ionic strength medium and release them if the pH and ionic strength are raised. Botulinum toxin was eluted with lower ionic strength than tetanus toxin, and could be freed from nontoxic admixtures. Analysis by affinity chromatography disclosed partially toxoided tetanus toxin in both labelled and unlabelled toxin samples. High concentrations of formaldehyde (0.5%) destroyed both toxicity and affinity to the synaptosomes of tetanus toxin. Low concentrations of formaldehyde (0.05%) yielded a derivative of low toxicity which was still, however less firmly, bound to synaptosomes. Tetanus and botulinum toxin differ by their acceptors. Whereas unlabelled botulinum toxin is unable to compete with labelled tetanus toxin, unlabelled tetanus toxin slightly competes with botulinum toxin. Both labelled toxins display anomalous binding behaviour in that they cannot be displaced completely even with a large excess of unlabelled toxin. (orig.) [de

  20. Vaccination of pregnant cows with EspA, EspB, γ-intimin, and Shiga toxin 2 proteins from Escherichia coli O157:H7 induces high levels of specific colostral antibodies that are transferred to newborn calves.

    Science.gov (United States)

    Rabinovitz, B C; Gerhardt, E; Tironi Farinati, C; Abdala, A; Galarza, R; Vilte, D A; Ibarra, C; Cataldi, A; Mercado, E C

    2012-06-01

    Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a major cause of intestinal disease and hemolytic uremic syndrome, a serious systemic complication that particularly affects children. Cattle are primary reservoirs for EHEC O157:H7 and the main source of infection for humans. Vaccination of cattle with different combinations of bacterial virulence factors has shown efficacy in decreasing EHEC O157:H7 shedding. It is, therefore, important to demonstrate whether vaccination of pregnant cows with EHEC O157:H7 induces high titers of transferable antibodies to avoid early colonization of calves by the bacteria. In this study we evaluated the ability of EspA, EspB, the C-terminal fragment of 280 amino acids of γ-intimin (γ-intimin C₂₈₀) and inactivated Shiga toxin (Stx) 2 proteins to induce specific antibodies in colostrum and their passive transference to colostrum-fed calves. Friesian pregnant cows immunized by the intramuscular route mounted significantly high serum and colostrum IgG responses against EspB and γ-intimin C₂₈₀ that were efficiently transferred to their calves. Antibodies to EspB and γ-intimin C₂₈₀ were detected in milk samples of vaccinated cows at d 40 postparturition. Significant Stx2-neutralizing titers were also observed in colostrum from Stx2-vaccinated cows and sera from colostrum-fed calves. The results presented showed that bovine colostrum with increased levels of antibodies against EHEC O157:H7 may be obtained by systemic immunization of pregnant cows, and that these specific antibodies are efficiently transferred to newborn calves by feeding colostrum. Hyperimmune colostrum and milk may be an alternative to protect calves from early colonization by EHEC O157:H7 and a possible key source of antibodies to block colonization and toxic activity of this bacterium. Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  1. Sequence variation in the alpha-toxin encoding plc gene of Clostridium perfringens strains isolated from diseased and healthy chickens

    DEFF Research Database (Denmark)

    Abildgaard, L; Engberg, RM; Pedersen, Karl

    2009-01-01

    The aim of the present study was to analyse the genetic diversity of the alpha-toxin encoding plc gene and the variation in a-toxin production of Clostridium perfringens type A strains isolated from presumably healthy chickens and chickens suffering from either necrotic enteritis (NE) or cholangio......-hepatitis. The a-toxin encoding plc genes from 60 different pulsed-field gel electrophoresis (PFGE) types (strains) of C perfringens were sequenced and translated in silico to amino acid sequences and the a-toxin production was investigated in batch cultures of 45 of the strains using an enzyme...

  2. Regulating Toxin-Antitoxin Expression: Controlled Detonation of Intracellular Molecular Timebombs

    Directory of Open Access Journals (Sweden)

    Finbarr Hayes

    2014-01-01

    Full Text Available Genes for toxin-antitoxin (TA complexes are widely disseminated in bacteria, including in pathogenic and antibiotic resistant species. The toxins are liberated from association with the cognate antitoxins by certain physiological triggers to impair vital cellular functions. TAs also are implicated in antibiotic persistence, biofilm formation, and bacteriophage resistance. Among the ever increasing number of TA modules that have been identified, the most numerous are complexes in which both toxin and antitoxin are proteins. Transcriptional autoregulation of the operons encoding these complexes is key to ensuring balanced TA production and to prevent inadvertent toxin release. Control typically is exerted by binding of the antitoxin to regulatory sequences upstream of the operons. The toxin protein commonly works as a transcriptional corepressor that remodels and stabilizes the antitoxin. However, there are notable exceptions to this paradigm. Moreover, it is becoming clear that TA complexes often form one strand in an interconnected web of stress responses suggesting that their transcriptional regulation may prove to be more intricate than currently understood. Furthermore, interference with TA gene transcriptional autoregulation holds considerable promise as a novel antibacterial strategy: artificial release of the toxin factor using designer drugs is a potential approach to induce bacterial suicide from within.

  3. Methods for detecting pathogens in the beef food chain: detecting particular pathogens

    Science.gov (United States)

    The main food-borne pathogens of concern in the beef food chain are Shiga toxin-producing Escherichia coli (STEC) and Salmonella spp.; however, the presence of other pathogens, including Listeria monocytogenes, Campylobacter spp., Clostridium spp., Bacillus cereus, and Mycobacterium avium subsp. par...

  4. Toxin-Based Therapeutic Approaches

    Directory of Open Access Journals (Sweden)

    Itai Benhar

    2010-10-01

    Full Text Available Protein toxins confer a defense against predation/grazing or a superior pathogenic competence upon the producing organism. Such toxins have been perfected through evolution in poisonous animals/plants and pathogenic bacteria. Over the past five decades, a lot of effort has been invested in studying their mechanism of action, the way they contribute to pathogenicity and in the development of antidotes that neutralize their action. In parallel, many research groups turned to explore the pharmaceutical potential of such toxins when they are used to efficiently impair essential cellular processes and/or damage the integrity of their target cells. The following review summarizes major advances in the field of toxin based therapeutics and offers a comprehensive description of the mode of action of each applied toxin.

  5. Staphylococcal β-Toxin Modulates Human Aortic Endothelial Cell and Platelet Function through Sphingomyelinase and Biofilm Ligase Activities

    Directory of Open Access Journals (Sweden)

    Alfa Herrera

    2017-03-01

    Full Text Available Staphylococcus aureus causes many infections, such as skin and soft tissue, pneumonia, osteomyelitis, and infective endocarditis (IE. IE is an endovascular infection of native and prosthetic valves and the lining of the heart; it is characterized by the formation of cauliflower-like “vegetations” composed of fibrin, platelets, other host factors, bacteria, and bacterial products. β-Toxin is an S. aureus virulence factor that contributes to the microorganism’s ability to cause IE. This cytolysin has two enzymatic activities: sphingomyelinase (SMase and biofilm ligase. Although both activities have functions in a rabbit model of IE, the mechanism(s by which β-toxin directly affects human cells and is involved in the infectious process has not been elucidated. Here, we compared the in vitro effects of purified recombinant wild-type β-toxin, SMase-deficient β-toxin (H289N, and biofilm ligase-deficient β-toxin (H162A and/or D163A on human aortic endothelial cells (HAECs and platelets. β-Toxin was cytotoxic to HAECs and inhibited the production of interleukin 8 (IL-8 from these cells by both SMase and biofilm ligase activities. β-Toxin altered HAEC surface expression of CD40 and vascular cell adhesion molecule 1 (VCAM-1. HAECs treated with β-toxin displayed granular membrane morphology not seen in treatment with the SMase-deficient mutant. The altered morphology resulted in two possibly separable activities, cell rounding and redistribution of cell membranes into granules, which were not the result of endosome production from the Golgi apparatus or lysosomes. β-Toxin directly aggregated rabbit platelets via SMase activity.

  6. Deoxynivalenol and other Fusarium toxins in wheat and rye flours on the Danish market

    DEFF Research Database (Denmark)

    Rasmussen, Peter Have; Pasikhani, Faranak Ghorbani; Berg, T.

    2003-01-01

    Information on the contamination of Danish cereals and cereal products with Fusarium toxins is limited and the last survey is from 1984/1985. In the present study, the occurrence of deoxynivalenol (DON), nivalenol (NIV), HT-2 toxin, T-2 toxin and zearalenone (ZON) was investigated in. our of common...... wheat, durum wheat and rye. The samples were collected from 1998 to 2001 from both mills and the retail market in Denmark. A total of 190. our samples were analysed for DON and NIV and about 60 samples for HT-2, T-2 toxin and ZON. DON was most frequently detected with an incidence rate of 78% over all......)). Contents of NIV, HT-2 toxin and ZON in samples of wheat and rye were generally low, and even in positive samples the contents were close to the detection limit of the methods. The T-2 toxin was detected in only a few of the wheat samples and in low amounts. However, the toxin was found in about 50...

  7. Bioengineered kidney tubules efficiently excrete uremic toxins

    NARCIS (Netherlands)

    Jansen, Jitske; Fedecostante, M.; Wilmer, M.; Peters, J.G.; Kreuser, U.M.; Broek, P.H.; Mensink, R.A.; Boltje, T.J.; Stamatialis, Dimitrios; Wetzels, J.F.; van der Heuvel, L.P.; Hoenderop, J.G.; Masereeuw, R.

    2016-01-01

    The development of a biotechnological platform for the removal of waste products (e.g. uremic toxins), often bound to proteins in plasma, is a prerequisite to improve current treatment modalities for patients suffering from end stage renal disease (ESRD). Here, we present a newly designed

  8. Epidermal Growth Factor Receptor Signaling Enhances the Proinflammatory Effects of Staphylococcus aureus Gamma-Toxin on the Mucosa.

    Science.gov (United States)

    Gillman, Aaron N; Breshears, Laura M; Kistler, Charles K; Finnegan, Patrick M; Torres, Victor J; Schlievert, Patrick M; Peterson, Marnie L

    2017-06-28

    Staphylococcus aureus ( S. aureus ) produces many different exotoxins including the gamma-toxins, HlgAB and HlgCB. Gamma-toxins form pores in both leukocyte and erythrocyte membranes, resulting in cell lysis. The genes encoding gamma-toxins are present in most strains of S. aureus, and are commonly expressed in clinical isolates recovered from menstrual Toxic Shock Syndrome (mTSS) patients. This study set out to investigate the cytotoxic and proinflammatory effects of gamma-toxins on vaginal epithelial surfaces. We found that both HlgAB and HlgCB were cytotoxic to cultured human vaginal epithelial cells (HVECs) and induced cytokine production at sub-cytotoxic doses. Cytokine production induced by gamma-toxin treatment of HVECs was found to involve epidermal growth factor receptor (EGFR) signaling and mediated by shedding of EGFR ligands from the cell surface. The gamma-toxin subunits displayed differential binding to HVECs (HlgA 93%, HlgB 97% and HlgC 28%) with both components (HlgAB or HlgCB) required for maximum detectable binding and significant stimulation of cytokine production. In studies using full thickness ex vivo porcine vaginal mucosa, HlgAB or HlgCB stimulated a dose-dependent cytokine response, which was reduced significantly by inhibition of EGFR signaling. The effects of gamma-toxins on porcine vaginal tissue and cultured HVECs were validated using ex vivo human ectocervical tissue. Collectively, these studies have identified the EGFR-signaling pathway as a key component in gamma-toxin-induced proinflammatory changes at epithelial surfaces and highlight a potential therapeutic target to diminish toxigenic effects of S. aureus infections.

  9. Nanomaterial-based sensors for detection of foodborne bacterial pathogens and toxins as well as pork adulteration in meat products

    Directory of Open Access Journals (Sweden)

    B. Stephen Inbaraj

    2016-01-01

    Full Text Available Food safety draws considerable attention in the modern pace of the world owing to rapid-changing food recipes and food habits. Foodborne illnesses associated with pathogens, toxins, and other contaminants pose serious threat to human health. Besides, a large amount of money is spent on both analyses and control measures, which causes significant loss to the food industry. Conventional detection methods for bacterial pathogens and toxins are time consuming and laborious, requiring certain sophisticated instruments and trained personnel. In recent years, nanotechnology has emerged as a promising field for solving food safety issues in terms of detecting contaminants, enabling controlled release of preservatives to extend the shelf life of foods, and improving food-packaging strategies. Nanomaterials including metal oxide and metal nanoparticles, carbon nanotubes, and quantum dots are gaining a prominent role in the design of sensors and biosensors for food analysis. In this review, various nanomaterial-based sensors reported in the literature for detection of several foodborne bacterial pathogens and toxins are summarized highlighting their principles, advantages, and limitations in terms of simplicity, sensitivity, and multiplexing capability. In addition, the application through a noncross-linking method without the need for any surface modification is also presented for detection of pork adulteration in meat products.

  10. The effect of chronic feeding of diacetoxyscirpenol and T-2 toxin on performance, health, small intestinal physiology and antibody production in turkey poults.

    Science.gov (United States)

    Sklan, D; Shelly, M; Makovsky, B; Geyra, A; Klipper, E; Friedman, A

    2003-03-01

    1. The effects of feeding T-2 toxin or diacetoxyscirpenol (DAS) at levels up to 1 ppm for 32 d on performance, health, small intestinal physiology and immune response to enteral and parenteral immunisation were examined in young poults. 2. Slight improvement in growth was observed in some groups of poults fed T-2 or DAS mycotoxins for 32 d, with no change in feed efficiency. Feeding both T-2 and DAS resulted in oral lesions which had maximal severity after 7-15 d. 3. Mild intestinal changes were observed at 32 d but no pathological or histopathological lesions were found. Both mycotoxins altered small intestinal morphology, especially in the jejunum where villi were shorter and thinner. In addition, both DAS and T-2 mycotoxins enhanced the proportion of proliferating cells both in the crypts and along the villi. Migration rates were reduced in the jejunum of poults fed T-2 toxin but did not change in the duodenum or in poults fed DAS. 4. No significant effects of T-2 or DAS were observed on antibody production to antigens administered by enteral or parenteral routes. 5. This study indicates that tricothecene toxins at concentrations of up to 1 ppm for more than 30 d influenced small intestinal morphology but did not affect growth or antibody production.

  11. From lin-Benzoguanines to lin-Benzohypoxanthines as Ligands for Zymomonas mobilis tRNA-Guanine Transglycosylase: Replacement of Protein-Ligand Hydrogen Bonding by Importing Water Clusters.

    NARCIS (Netherlands)

    Barandun, L.J.; Immekus, F.; Kohler, P.C.; Tonazzi, S.; Wagner, B.; Wendelspiess, S.; Ritschel, T.; Heine, A.; Kansy, M.; Klebe, G.; Diederich, F.

    2012-01-01

    The foodborne illness shigellosis is caused by Shigella bacteria that secrete the highly cytotoxic Shiga toxin, which is also formed by the closely related enterohemorrhagic Escherichia coli (EHEC). It has been shown that tRNA-guanine transglycosylase (TGT) is essential for the pathogenicity of

  12. Microbiota response to Escherichia coli O157:H7 colonization in cattle

    Science.gov (United States)

    Cattle are primary reservoir of Shiga toxin-producing Escherichia coli (STEC). Field studies indicate STEC colonization influences gut microbiota composition in beef and dairy cattle. In this pilot study, we evaluated the bovine gut microbiota after STEC O157 (O157) challenge under experimental con...

  13. Botulinum Toxin (Botox) for Facial Wrinkles

    Science.gov (United States)

    ... Stories Español Eye Health / Eye Health A-Z Botulinum Toxin (Botox) for Facial Wrinkles Sections Botulinum Toxin (Botox) ... Facial Wrinkles How Does Botulinum Toxin (Botox) Work? Botulinum Toxin (Botox) for Facial Wrinkles Leer en Español: La ...

  14. Veal calves produce less antibodies against C. perfringens alpha toxin compared to beef calves

    OpenAIRE

    Valgaeren, Bonnie; Pardon, Bart; Goossens, Evy; Verherstraeten, Stefanie; Roelandt, Sophie; Timbermont, Leen; Van Der Vekens, Nicky; Stuyvaert, Sabrina; Gille, Linde; Van Driessche, Laura; Haesebrouck, Freddy; Ducatelle, Richard; Van Immerseel, Filip; Deprez, Piet

    2015-01-01

    Enterotoxaemia is a disease with a high associated mortality rate, affecting beef and veal calves worldwide, caused by C. perfringens alpha toxin and perfringolysin. A longitudinal study was conducted to determine the dynamics of antibodies against these toxins in 528 calves on 4 beef and 15 veal farms. The second study aimed to determine the effect of solid feed intake on the production of antibodies against alpha toxin and perfringolysin. The control group only received milk replacer, wher...

  15. Lymphocyte receptors for pertussis toxin

    Energy Technology Data Exchange (ETDEWEB)

    Clark, C.G.; Armstrong, G.D. (Univ. of Alberta, Edmonton (Canada))

    1990-12-01

    We have investigated human T-lymphocyte receptors for pertussis toxin by affinity isolation and photoaffinity labeling procedures. T lymphocytes were obtained from peripheral human blood, surface iodinated, and solubilized in Triton X-100. The iodinated mixture was then passed through pertussis toxin-agarose, and the fractions were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Autoradiography of the fixed, dried gels revealed several bands in the pertussis toxin-bound fraction that were not observed in fractions obtained from histone or fetuin-agarose. Further investigations employed a photoaffinity labeling reagent, sulfosuccinimidyl 2-(p-azido-salicylamido)-1,3'-dithiopropionate, to identify pertussis toxin receptors in freshly isolated peripheral blood monocytic cells, T lymphocytes, and Jurkat cells. In all three cell systems, the pertussis toxin affinity probe specifically labeled a single protein species with an apparent molecular weight of 70,000 that was not observed when the procedure was performed in the presence of excess unmodified pertussis toxin. A protein comparable in molecular weight to the one detected by the photoaffinity labeling technique was also observed among the species that bound to pertussis toxin-agarose. The results suggest that pertussis toxin may bind to a 70,000-Da receptor in human T lymphocytes.

  16. Occurrence and elimination of cyanobacterial toxins in drinking water treatment plants

    International Nuclear Information System (INIS)

    Hoeger, Stefan J.; Hitzfeld, Bettina C.; Dietrich, Daniel R.

    2005-01-01

    Toxin-producing cyanobacteria (blue-green algae) are abundant in surface waters used as drinking water resources. The toxicity of one group of these toxins, the microcystins, and their presence in surface waters used for drinking water production has prompted the World Health Organization (WHO) to publish a provisional guideline value of 1.0 μg microcystin (MC)-LR/l drinking water. To verify the efficiency of two different water treatment systems with respect to reduction of cyanobacterial toxins, the concentrations of MC in water samples from surface waters and their associated water treatment plants in Switzerland and Germany were investigated. Toxin concentrations in samples from drinking water treatment plants ranged from below 1.0 μg MC-LR equiv./l to more than 8.0 μg/l in raw water and were distinctly below 1.0 μg/l after treatment. In addition, data to the worldwide occurrence of cyanobacteria in raw and final water of water works and the corresponding guidelines for cyanobacterial toxins in drinking water worldwide are summarized

  17. Topical botulinum toxin.

    Science.gov (United States)

    Collins, Ashley; Nasir, Adnan

    2010-03-01

    Nanotechnology is a rapidly growing discipline that capitalizes on the unique properties of matter engineered on the nanoscale. Vehicles incorporating nanotechnology have led to great strides in drug delivery, allowing for increased active ingredient stability, bioavailability, and site-specific targeting. Botulinum toxin has historically been used for the correction of neurological and neuromuscular disorders, such as torticollis, blepharospasm, and strabismus. Recent dermatological indications have been for the management of axillary hyperhydrosis and facial rhytides. Traditional methods of botulinum toxin delivery have been needle-based. These have been associated with increased pain and cost. Newer methods of botulinum toxin formulation have yielded topical preparations that are bioactive in small pilot clinical studies. While there are some risks associated with topical delivery, the refinement and standardization of delivery systems and techniques for the topical administration of botulinum toxin using nanotechnology is anticipated in the near future.

  18. Potency of a human monoclonal antibody to diphtheria toxin relative to equine diphtheria anti-toxin in a guinea pig intoxication model.

    Science.gov (United States)

    Smith, Heidi L; Cheslock, Peter; Leney, Mark; Barton, Bruce; Molrine, Deborah C

    2016-08-17

    Prompt administration of anti-toxin reduces mortality following Corynebacterium diphtheriae infection. Current treatment relies upon equine diphtheria anti-toxin (DAT), with a 10% risk of serum sickness and rarely anaphylaxis. The global DAT supply is extremely limited; most manufacturers have ceased production. S315 is a neutralizing human IgG1 monoclonal antibody to diphtheria toxin that may provide a safe and effective alternative to equine DAT and address critical supply issues. To guide dose selection for IND-enabling pharmacology and toxicology studies, we dose-ranged S315 and DAT in a guinea pig model of diphtheria intoxication based on the NIH Minimum Requirements potency assay. Animals received a single injection of antibody premixed with toxin, were monitored for 30 days, and assigned a numeric score for clinical signs of disease. Animals receiving ≥ 27.5 µg of S315 or ≥ 1.75 IU of DAT survived whereas animals receiving ≤ 22.5 µg of S315 or ≤ 1.25 IU of DAT died, yielding a potency estimate of 17 µg S315/IU DAT (95% CI 16-21) for an endpoint of survival. Because some surviving animals exhibited transient limb weakness, likely a systemic sign of toxicity, DAT and S315 doses required to prevent hind limb paralysis were also determined, yielding a relative potency of 48 µg/IU (95% CI 38-59) for this alternate endpoint. To support advancement of S315 into clinical trials, potency estimates will be used to evaluate the efficacy of S315 versus DAT in an animal model with antibody administration after toxin exposure, more closely modeling anti-toxin therapy in humans.

  19. Diffusion, spread, and migration of botulinum toxin.

    Science.gov (United States)

    Ramirez-Castaneda, Juan; Jankovic, Joseph; Comella, Cynthia; Dashtipour, Khashayar; Fernandez, Hubert H; Mari, Zoltan

    2013-11-01

    Botulinum toxin (BoNT) is an acetylcholine release inhibitor and a neuromuscular blocking agent used for the treatment of a variety of neurologic and medical conditions. The efficacy and safety of BoNT depends on accurate selection and identification of intended targets but also may be determined by other factors, including physical spread of the molecule from the injection site, passive diffusion, and migration to distal sites via axonal or hematogenous transport. The passive kinetic dispersion of the toxin away from the injection site in a gradient-dependent manner may also play a role in toxin spread. In addition to unique properties of the various BoNT products, volume and dilution may also influence local and systemic distribution of BoNT. Most of the local and remote complications of BoNT injections are thought to be due to unwanted spread or diffusion of the toxin's biologic activity into adjacent and distal muscles. Despite widespread therapeutic and cosmetic use of BoNT over more than three decades, there is a remarkable paucity of published data on the mechanisms of distribution and its effects on clinical outcomes. The primary aim of this article is to critically review the available experimental and clinical literature and place it in the practical context. © 2013 International Parkinson and Movement Disorder Society.

  20. Tetrodotoxin-Producing Bacteria: Detection, Distribution and Migration of the Toxin in Aquatic Systems

    Directory of Open Access Journals (Sweden)

    Timur Yu. Magarlamov

    2017-05-01

    Full Text Available This review is devoted to the marine bacterial producers of tetrodotoxin (TTX, a potent non-protein neuroparalytic toxin. In addition to the issues of the ecology and distribution of TTX-producing bacteria, this review examines issues relating to toxin migration from bacteria to TTX-bearing animals. It is shown that the mechanism of TTX extraction from toxin-producing bacteria to the environment occur through cell death, passive/active toxin excretion, or spore germination of spore-forming bacteria. Data on TTX microdistribution in toxic organs of TTX-bearing animals indicate toxin migration from the digestive system to target organs through the transport system of the organism. The role of symbiotic microflora in animal toxicity is also discussed: despite low toxin production by bacterial strains in laboratory conditions, even minimal amounts of TTX produced by intestinal microflora of an animal can contribute to its toxicity. Special attention is paid to methods of TTX detection applicable to bacteria. Due to the complexity of toxin detection in TTX-producing bacteria, it is necessary to use several methods based on different methodological approaches. Issues crucial for further progress in detecting natural sources of TTX investigation are also considered.

  1. Botulinum toxin injection - larynx

    Science.gov (United States)

    Injection laryngoplasty; Botox - larynx: spasmodic dysphonia-BTX; Essential voice tremor (EVT)-btx; Glottic insufficiency; Percutaneous electromyography - guided botulinum toxin treatment; Percutaneous indirect laryngoscopy - guided botulinum toxin treatment; ...

  2. Standardization of the PCR technique for the detection of delta toxin in Staphylococcus spp.

    Directory of Open Access Journals (Sweden)

    C. Marconi

    2005-06-01

    Full Text Available Coagulase-negative staphylococci (CNS, components of the normal flora of neonates, have emerged as important opportunistic pathogens of nosocomial infections that occur in neonatal intensive care units. Some authors have reported the ability of some CNS strains, particularly Staphylococcus epidermidis, to produce a toxin similar to S. aureus delta toxin. This toxin is an exoprotein that has a detergent action on the membranes of various cell types resulting in rapid cell lysis. The objectives of the present study were to standardize the Polymerase Chain Reaction (PCR technique for the detection of the gene responsible for the production of delta toxin (hld gene in staphylococcal species isolated from catheters and blood cultures obtained from neonates, and to compare the results to those obtained with the phenotypic synergistic hemolysis method. Detection of delta toxin by the phenotypic and genotypic method yielded similar results for the S. aureus isolates. However, in S. epidermidis, a higher positivity was observed for PCR (97.4% compared to the synergistic hemolysis method (86.8%. Among CNS, S. epidermidis was the most frequent isolate and was a delta toxin producer. Staphylococcus simulans and S. warneri tested positive by the phenotypic method, but their positivity was not confirmed by PCR for the hld gene detection. These results indicate that different genes might be responsible for the production of this toxin in different CNS species, requiring highly specific primers for their detection. PCR was found to be a rapid and reliable method for the detection of the hld gene in S. aureus and S. epidermidis.

  3. Verocytotoxin inhibits mitogenesis and protein synthesis in purified human glomerular mesangial cells without affecting cell viability: Evidence for two distinct mechanisms

    NARCIS (Netherlands)

    Setten, P.A. van; Hinsbergh, V.W.M. van; Heuvel, L.P.W.J. van den; Velden, T.J.A.N. van der; Kar, N.C.A.J. van de; Krebbers, R.J.M.; Karmali, M.A.; Monnens, L.A.H.

    1997-01-01

    Acute renal failure is one of the hallmarks of the hemolytic uremic syndrome (HUS). Infection with a verocytotoxin (VT)- or Shiga-like toxin (SLT)-producing Escherichia coli has been strongly implicated in the etiology of the epidemic form of HUS. The functional receptor for these closely related

  4. Complete Genome Sequence of an Escherichia coli O121:H19 Strain from an Outbreak in Canada Associated with Flour

    Science.gov (United States)

    Robertson, James; Lin, Janet; Levett, Paul N.; Nadon, Celine; Nash, John

    2018-01-01

    ABSTRACT Here, we present the first complete genome sequence of an Escherichia coli non-O157 Shiga-toxin producing isolate, 16-9255, from serotype O121:H19. This strain is notable as a clinical case recovered from a recent Canadian flour-associated outbreak event. PMID:29371368

  5. 75 FR 81613 - Proposed Data Collections Submitted for Public Comment and Recommendations

    Science.gov (United States)

    2010-12-28

    ...-O157 Shiga Toxin-Producing E. coli Study: Assessment of Risk Factors for Laboratory-Confirmed... DEPARTMENT OF HEALTH AND HUMAN SERVICES Centers for Disease Control and Prevention [60Day-11-11BI... States, ranging in severity from mild diarrhea, to hemorrhagic colitis and in some cases, life...

  6. Hemolytic Uremic Syndrome: New Developments in Pathogenesis and Treatment

    Directory of Open Access Journals (Sweden)

    Olivia Boyer

    2011-01-01

    Full Text Available Hemolytic uremic syndrome is defined by the characteristic triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. In children, most cases of HUS are caused by Shiga-toxin-producing bacteria, especially Escherichia coli O157:H7. Common vehicles of transmission include ground beef, unpasteurized milk, and municipal or swimming water. Shiga-toxin-associated HUS is a main cause of acute renal failure in young children. Management remains supportive as there is at present no specific therapy to ameliorate the prognosis. Immediate outcome is most often favourable but long-term renal sequelae are frequent due to nephron loss. Atypical HUS represents 5% of cases. In the past 15 years, mutations in complement regulators of the alternative pathway have been identified in almost 60% of cases, leading to excessive complement activation. The disease has a relapsing course and more than half of the patients either die or progress to end-stage renal failure. Recurrence after renal transplantation is frequent.

  7. Investigation of an outbreak of bloody diarrhea complicated with hemolytic uremic syndrome

    Directory of Open Access Journals (Sweden)

    Otar Chokoshvili

    2014-12-01

    Full Text Available In July–August 2009, eight patients with bloody diarrhea complicated by hemolytic uremic syndrome (HUS were admitted to hospitals in Tbilisi, Georgia. We started active surveillance in two regions for bloody diarrhea and post-diarrheal HUS. Of 25 case-patients who developed HUS, including the initial 8 cases, half were ⩾15 years old, 67% were female and seven (28% died. No common exposures were identified. Among 20 HUS case-patients tested, Shiga toxin was detected in the stools of 2 patients (one with elevated serum IgG titers to several Escherichia coli serogroups, including O111 and O104. Among 56 persons with only bloody diarrhea, we isolated Shiga toxin-producing E. coli (STEC O104:H4 from 2 and Shigella from 10; 2 had serologic evidence of E. coli O26 infection. These cases may indicate a previously unrecognized burden of HUS in Georgia. We recommend national reporting of HUS and improving STEC detection capacity.

  8. Development of a recombinant toxin fragment vaccine for Clostridium difficile infection.

    Science.gov (United States)

    Karczewski, Jerzy; Zorman, Julie; Wang, Su; Miezeiewski, Matthew; Xie, Jinfu; Soring, Keri; Petrescu, Ioan; Rogers, Irene; Thiriot, David S; Cook, James C; Chamberlin, Mihaela; Xoconostle, Rachel F; Nahas, Debbie D; Joyce, Joseph G; Bodmer, Jean-Luc; Heinrichs, Jon H; Secore, Susan

    2014-05-19

    Clostridium difficile infection (CDI) is the major cause of antibiotic-associated diarrhea and pseudomembranous colitis, a disease associated with significant morbidity and mortality. The disease is mostly of nosocomial origin, with elderly patients undergoing anti-microbial therapy being particularly at risk. C. difficile produces two large toxins: Toxin A (TcdA) and Toxin B (TcdB). The two toxins act synergistically to damage and impair the colonic epithelium, and are primarily responsible for the pathogenesis associated with CDI. The feasibility of toxin-based vaccination against C. difficile is being vigorously investigated. A vaccine based on formaldehyde-inactivated Toxin A and Toxin B (toxoids) was reported to be safe and immunogenic in healthy volunteers and is now undergoing evaluation in clinical efficacy trials. In order to eliminate cytotoxic effects, a chemical inactivation step must be included in the manufacturing process of this toxin-based vaccine. In addition, the large-scale production of highly toxic antigens could be a challenging and costly process. Vaccines based on non-toxic fragments of genetically engineered versions of the toxins alleviate most of these limitations. We have evaluated a vaccine assembled from two recombinant fragments of TcdB and explored their potential as components of a novel experimental vaccine against CDI. Golden Syrian hamsters vaccinated with recombinant fragments of TcdB combined with full length TcdA (Toxoid A) developed high titer IgG responses and potent neutralizing antibody titers. We also show here that the recombinant vaccine protected animals against lethal challenge with C. difficile spores, with efficacy equivalent to the toxoid vaccine. The development of a two-segment recombinant vaccine could provide several advantages over toxoid TcdA/TcdB such as improvements in manufacturability. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. [Intoxication of botulinum toxin].

    Science.gov (United States)

    Chudzicka, Aleksandra

    2015-09-01

    Botulinum toxin is an egzotoxin produced by Gram positive bacteria Clostridium botulinum. It is among the most potent toxins known. The 3 main clinical presentations of botulism are as follows: foodborne botulism, infant botulism and wound botulism. The main symptom of intoxication is flat muscles paralysis. The treatment is supportive care and administration of antitoxin. In prevention the correct preparing of canned food is most important. Botulinum toxin is accepted as a biological weapon. © 2015 MEDPRESS.

  10. Why do we study animal toxins?

    Science.gov (United States)

    ZHANG, Yun

    2015-01-01

    Venom (toxins) is an important trait evolved along the evolutionary tree of animals. Our knowledges on venoms, such as their origins and loss, the biological relevance and the coevolutionary patterns with other organisms are greatly helpful in understanding many fundamental biological questions, i.e., the environmental adaptation and survival competition, the evolution shaped development and balance of venoms, and the sophisticated correlations among venom, immunity, body power, intelligence, their genetic basis, inherent association, as well as the cost-benefit and trade-offs of biological economy. Lethal animal envenomation can be found worldwide. However, from foe to friend, toxin studies have led lots of important discoveries and exciting avenues in deciphering and fighting human diseases, including the works awarded the Nobel Prize and lots of key clinic therapeutics. According to our survey, so far, only less than 0.1% of the toxins of the venomous animals in China have been explored. We emphasize on the similarities shared by venom and immune systems, as well as the studies of toxin knowledge-based physiological toxin-like proteins/peptides (TLPs). We propose the natural pairing hypothesis. Evolution links toxins with humans. Our mission is to find out the right natural pairings and interactions of our body elements with toxins, and with endogenous toxin-like molecules. Although, in nature, toxins may endanger human lives, but from a philosophical point of view, knowing them well is an effective way to better understand ourselves. So, this is why we study toxins. PMID:26228472

  11. FimH has a strain and host-cell type dependent role in adherence of E. coli O157:H7 super-shedder strains to host cells

    Science.gov (United States)

    Escherichia coli O157:H7 (O157) are Shiga toxin-producing food-borne pathogens that are a significant threat to human health, causing severe illnesses including hemorrhagic uremic syndrome and kidney failure. Cattle are the major reservoirs of O157, with asymptomatic animals harboring the organism i...

  12. 76 FR 13191 - Agency Forms Undergoing Paperwork Reduction Act Review

    Science.gov (United States)

    2011-03-10

    .... Proposed Project FoodNet Non-O157 Shiga toxin-Producing E. coli Study: Assessment of Risk Factors for... DEPARTMENT OF HEALTH AND HUMAN SERVICES Centers for Disease Control and Prevention [30-Day-11-11BI... cases, life-threatening hemolytic uremic syndrome (HUS). HUS occurs most frequently following infection...

  13. Prevention, control and detection of Fusarial toxins

    Directory of Open Access Journals (Sweden)

    Nešić Ksenija D.

    2013-01-01

    Full Text Available The past couple of decades have provided considerable details on fungi and the toxins that they produce, as well on the mechanism of toxin action, toxicity and effects on animal and human health. But, since they are natural contaminants, their presence is often inevitable. Fusaria are widespread in all cereal-growing territories of the world, but they are especially common in our geographic area. Therefore, special attention is paid to the prevention and control, and also to the improvement of methods for their detection. Although all collected data were critical for understanding this worldwide problem, managing the impact of these toxins on the feed and food safety is still great practical challenge. There are a number of approaches that can be taken to minimize mycotoxin contamination in this chain: prevention of fungal growth and thus mycotoxin formation, strategies to reduce or eliminate mycotoxins from contaminated feedstuffs or diverting the contaminated products to low risk uses. A control program for mycotoxins from field to table should in­volve the criteria of an HACCP (Hazard Analysis Critical Control Points approach. It requires an understanding of the important aspects of the interactions of the toxigenic fungi with crop plants, the on-farm production and harvest methods for crops, the production of livestock using grains and processed feeds, including diagnostic capabilities for mycotoxicoses, and all the way to the development of processed foods for human consumption, as well as understanding the marketing and trade channels including storage and delivery of foods to the consumer’s table. A good testing protocol for mycotoxins is necessary to manage all of the control points and in order to be able to ensure a food supply free of toxic levels of mycotoxins for the consumer. [Projekat Ministarstva nauke Republike Srbije, br. III 46009

  14. Strategies for On-Line Decontamination of Carcasses

    Science.gov (United States)

    Byelashov, Oleksandr A.; Sofos, John N.

    Microbial food safety has been one of the most important challenges for the meat industry during the last two decades due to important foodborne outbreaks traced to contaminated products and associated costly product recalls from the market. Escherichia coli O157:H7 and other non-O157 Shiga toxin-producing (STEC) strains, as well as Salmonella serotypes, Campylobacter jejuni, Clostridium perfringens, Clostridium botulinum, Listeria monocytogenes, Staphylococcus aureus, Yersinia enterocolitica, Aeromonas hydrophila, and Bacillus cereusare important pathogenic contaminants of meat and poultry products (Sofos, 2004a). STEC, especially, have been of major concern for the beef industry for a number of years, since for almost two decades contaminated beef products have been major sources of foodborne E. coli O157:H7 infection (Rangel, Sparling, Crowe, Griffin, & Swerdlow, 2005).

  15. Investigation of inactivation of Clostridium botulinum toxin by nuclear radiation. Final report. Untersuchung zur Desaktivierung des Clostridium botulinum Toxins durch Kernstrahlung. Endbericht

    Energy Technology Data Exchange (ETDEWEB)

    Kaltenhaeuser, A.; Werner, K.H.

    1989-01-01

    The effect of nuclear radiation on the toxicity and the molecular structure of the toxin produced by the microorganism Clostridium botulinum type A was investigated. The radiation induced changes in the structure of the toxin molecule. This effect is influenced by the composition or the medium above the toxin solution as well as by the temperature during the irradiation. The results of the investigation indicate that with increasing irradiation dose a new molecule was formed with immunological properties similar to the properties of the original molecule however with a greater molecular weight. After exposure to a radiation dose of 3,4 Mrad at normal temperature in air, complete detoxification of the substance was found. Immunizing experiments with the toxoid with two guinea-pigs indicated a pronounced increase of the antibody titer in the serum after 4 weeks. Vaccination experiments with the toxoid on animals show, that the protection against the effect of the toxin corresponds to the demands of the European Pharmacopoeia. The efficiency of the toxoid shows a similar efficiency as toxoids produced by chemical methods. The production of a toxoid-viccine with the relatively simple method of nuclear radiation appears possible. (orig./MG) With 12 refs., 3 tabs., 11 figs.

  16. Synthesis and biology of cyclic imine toxins, an emerging class of potent, globally distributed marine toxins.

    Science.gov (United States)

    Stivala, Craig E; Benoit, Evelyne; Aráoz, Rómulo; Servent, Denis; Novikov, Alexei; Molgó, Jordi; Zakarian, Armen

    2015-03-01

    From a small group of exotic compounds isolated only two decades ago, Cyclic Imine (CI) toxins have become a major class of marine toxins with global distribution. Their distinct chemical structure, biological mechanism of action, and intricate chemistry ensures that CI toxins will continue to be the subject of fascinating fundamental studies in the broad fields of chemistry, chemical biology, and toxicology. The worldwide occurrence of potent CI toxins in marine environments, their accumulation in shellfish, and chemical stability are important considerations in assessing risk factors for human health. This review article aims to provide an account of chemistry, biology, and toxicology of CI toxins from their discovery to the present day.

  17. Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins

    NARCIS (Netherlands)

    Mantzouki, Evanthia; Lürling, Miquel; Fastner, Jutta; de Senerpont Domis, Lisette; Wilk-Woźniak, Elżbieta; Koreivienė, Judita; Seelen, Laura; Teurlincx, Sven; Verstijnen, Yvon; Krztoń, Wojciech; Walusiak, Edward; Karosienė, Jūratė; Kasperovičienė, Jūratė; Savadova, Ksenija; Vitonytė, Irma; Cillero-Castro, Carmen; Budzyńska, Agnieszka; Goldyn, Ryszard; Kozak, Anna; Rosińska, Joanna; Szeląg-Wasielewska, Elżbieta; Domek, Piotr; Jakubowska-Krepska, Natalia; Kwasizur, Kinga; Messyasz, Beata; Pełechaty, Aleksandra; Pełechaty, Mariusz; Kokocinski, Mikolaj; García-Murcia, Ana; Real, Monserrat; Romans, Elvira; Noguero-Ribes, Jordi; Duque, David Parreño; Fernández-Morán, Elísabeth; Karakaya, Nusret; Häggqvist, Kerstin; Demir, Nilsun; Beklioğlu, Meryem; Filiz, Nur; Levi, Eti E.; Iskin, Uğur; Bezirci, Gizem; Tavşanoğlu, Ülkü Nihan; Özhan, Koray; Gkelis, Spyros; Panou, Manthos; Fakioglu, Özden; Avagianos, Christos; Kaloudis, Triantafyllos; Çelik, Kemal; Yilmaz, Mete; Marcé, Rafael; Catalán, Nuria; Bravo, Andrea G.; Buck, Moritz; Colom-Montero, William; Mustonen, Kristiina; Pierson, Don; Yang, Yang; Raposeiro, Pedro M.; Gonçalves, Vítor; Antoniou, Maria G.; Tsiarta, Nikoletta; McCarthy, Valerie; Perello, Victor C.; Feldmann, Tõnu; Laas, Alo; Panksep, Kristel; Tuvikene, Lea; Gagala, Ilona; Mankiewicz-Boczek, Joana; Yağcı, Meral Apaydın; Çınar, Şakir; Çapkın, Kadir; Yağcı, Abdulkadir; Cesur, Mehmet; Bilgin, Fuat; Bulut, Cafer; Uysal, Rahmi; Obertegger, Ulrike; Boscaini, Adriano; Flaim, Giovanna; Salmaso, Nico; Cerasino, Leonardo; Richardson, Jessica; Visser, Petra M; Verspagen, Jolanda M. H.; Karan, Tünay; Soylu, Elif Neyran; Maraşlıoğlu, Faruk; Napiórkowska-Krzebietke, Agnieszka; Ochocka, Agnieszka; Pasztaleniec, Agnieszka; Antão-Geraldes, Ana M.; Vasconcelos, Vitor; Morais, João; Vale, Micaela; Köker, Latife; Akçaalan, Reyhan; Albay, Meriç; Špoljarić Maronić, Dubravka; Stević, Filip; Žuna Pfeiffer, Tanja; Fonvielle, Jeremy; Straile, Dietmar; Rothhaupt, Karl-Otto; Hansson, Lars-Anders; Urrutia-Cordero, Pablo; Bláha, Luděk; Geriš, Rodan; Fránková, Markéta; Koçer, Mehmet Ali Turan; Alp, Mehmet Tahir; Remec-Rekar, Spela; Elersek, Tina; Triantis, Theodoros; Zervou, Sevasti-Kiriaki; Hiskia, Anastasia; Haande, Sigrid; Skjelbred, Birger; Madrecka, Beata; Nemova, Hana; Drastichova, Iveta; Chomova, Lucia; Edwards, Christine; Sevindik, Tuğba Ongun; Tunca, Hatice; Önem, Burçin; Aleksovski, Boris; Krstić, Svetislav; Vucelić, Itana Bokan; Nawrocka, Lidia; Salmi, Pauliina; Machado-Vieira, Danielle; de Oliveira, Alinne Gurjão; Delgado-Martín, Jordi; García-García, David; Cereijo, Jose Luís; Gomà, Joan; Trapote, Mari Carmen; Vegas-Vilarrúbia, Teresa; Obrador, Biel; Grabowska, Magdalena; Karpowicz, Maciej; Chmura, Damian; Úbeda, Bárbara; Gálvez, José Ángel; Özen, Arda; Christoffersen, Kirsten Seestern; Warming, Trine Perlt; Kobos, Justyna; Mazur-Marzec, Hanna; Pérez-Martínez, Carmen; Ramos-Rodríguez, Eloísa; Arvola, Lauri; Alcaraz-Párraga, Pablo; Toporowska, Magdalena; Pawlik-Skowronska, Barbara; Niedźwiecki, Michał; Pęczuła, Wojciech; Leira, Manel; Hernández, Armand; Moreno-Ostos, Enrique; Blanco, José María; Rodríguez, Valeriano; Montes-Pérez, Jorge Juan; Palomino, Roberto L.; Rodríguez-Pérez, Estela; Carballeira, Rafael; Camacho, Antonio; Picazo, Antonio; Rochera, Carlos; Santamans, Anna C.; Ferriol, Carmen; Romo, Susana; Soria, Juan Miguel; Dunalska, Julita; Sieńska, Justyna; Szymański, Daniel; Kruk, Marek; Kostrzewska-Szlakowska, Iwona; Jasser, Iwona; Žutinić, Petar; Gligora Udovič, Marija; Plenković-Moraj, Anđelka; Frąk, Magdalena; Bańkowska-Sobczak, Agnieszka; Wasilewicz, Michał; Özkan, Korhan; Maliaka, Valentini; Kangro, Kersti; Grossart, Hans-Peter; Paerl, Hans W.; Carey, Cayelan C.; Ibelings, Bas W.

    2018-01-01

    Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins). Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a) and

  18. Role of dinoflagellatte associated bacteria in toxin production : application to Dinophysis spp and Prorocentrum lima

    OpenAIRE

    Rausch de Traunlenberg, C.

    1992-01-01

    Des travaux récents ont été réalisés sur le rôle des bactéries associées aux dinoflagellés. Ils tendent à montrer que les bactéries seraient impliquées dans la production de toxine, voire qu'elles en seraient les productrices. Aucun travail n'ayant à ce jour été réalisé sur les bactéries associées aux dinoflagellés responsables d'intoxication DSP, nous avons extrapolé ces données à ce type de dinoflagellé afin d'évaluer le rôle des bactéries associées à deux dinoflagellés producteurs de DSP ...

  19. Defense against Toxin Weapons

    National Research Council Canada - National Science Library

    Franz, David

    1998-01-01

    .... We typically fear what we do not understand. Although un- derstanding toxin poisoning is less useful in a toxin attack than knowledge of cold injury on an Arctic battlefield, information on any threat reduces its potential to harm...

  20. Food toxin detection with atomic force microscope

    Science.gov (United States)

    Externally introduced toxins or internal spoilage correlated pathogens and their metabolites are all potential sources of food toxins. To prevent and protect unsafe food, many food toxin detection techniques have been developed to detect various toxins for quality control. Although several routine m...

  1. Cellular Entry of Clostridium perfringens Iota-Toxin and Clostridium botulinum C2 Toxin.

    Science.gov (United States)

    Takehara, Masaya; Takagishi, Teruhisa; Seike, Soshi; Oda, Masataka; Sakaguchi, Yoshihiko; Hisatsune, Junzo; Ochi, Sadayuki; Kobayashi, Keiko; Nagahama, Masahiro

    2017-08-11

    Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin are composed of two non-linked proteins, one being the enzymatic component and the other being the binding/translocation component. These latter components recognize specific receptors and oligomerize in plasma membrane lipid-rafts, mediating the uptake of the enzymatic component into the cytosol. Enzymatic components induce actin cytoskeleton disorganization through the ADP-ribosylation of actin and are responsible for cell rounding and death. This review focuses upon the recent advances in cellular internalization of clostridial binary toxins.

  2. Conditional Toxin Splicing Using a Split Intein System.

    Science.gov (United States)

    Alford, Spencer C; O'Sullivan, Connor; Howard, Perry L

    2017-01-01

    Protein toxin splicing mediated by split inteins can be used as a strategy for conditional cell ablation. The approach requires artificial fragmentation of a potent protein toxin and tethering each toxin fragment to a split intein fragment. The toxin-intein fragments are, in turn, fused to dimerization domains, such that addition of a dimerizing agent reconstitutes the split intein. These chimeric toxin-intein fusions remain nontoxic until the dimerizer is added, resulting in activation of intein splicing and ligation of toxin fragments to form an active toxin. Considerations for the engineering and implementation of conditional toxin splicing (CTS) systems include: choice of toxin split site, split site (extein) chemistry, and temperature sensitivity. The following method outlines design criteria and implementation notes for CTS using a previously engineered system for splicing a toxin called sarcin, as well as for developing alternative CTS systems.

  3. Mutant with diphtheria toxin receptor and acidification function but defective in entry of toxin

    International Nuclear Information System (INIS)

    Kohno, Kenji; Hayes, H.; Mekada, Eisuke; Uchida, Tsuyoshi

    1987-01-01

    A mutant of Chinese hamster ovary cells, GE1, that is highly resistant to diphtheria toxin was isolated. The mutant contains 50% ADP-ribosylatable elongation factor 2, but its protein synthesis was not inhibited by the toxin even at concentrations above 100 μg/ml. 125 I-labeled diphtheria toxin was associated with GE1 cells as well as with the parent cells but did not block protein synthesis of GE1 cells even when the cells were exposed to low pH in the presence or absence of NH 4 Cl. The infections of GE1 cells and the parent cells by vesicular stomatitis virus were similar. GE1 cells were cross-resistant to Pseudomonas aeruginosa exotoxin A and so were about 1,000 times more resistant to this toxin than the parent cells. Hybrids of GE1 cells and the parent cells or mutant cells lacking a functional receptor were more sensitive to diphtheria toxin than GE1 cells. These results suggest that entry of diphtheria toxin into cells requires a cellular factor(s) in addition to those involved in receptor function and acidification of endosomes and that GE1 cells do not express this cellular factor. This character is recessive in GE1 cells

  4. Computational Studies of Snake Venom Toxins.

    Science.gov (United States)

    Ojeda, Paola G; Ramírez, David; Alzate-Morales, Jans; Caballero, Julio; Kaas, Quentin; González, Wendy

    2017-12-22

    Most snake venom toxins are proteins, and participate to envenomation through a diverse array of bioactivities, such as bleeding, inflammation, and pain, cytotoxic, cardiotoxic or neurotoxic effects. The venom of a single snake species contains hundreds of toxins, and the venoms of the 725 species of venomous snakes represent a large pool of potentially bioactive proteins. Despite considerable discovery efforts, most of the snake venom toxins are still uncharacterized. Modern bioinformatics tools have been recently developed to mine snake venoms, helping focus experimental research on the most potentially interesting toxins. Some computational techniques predict toxin molecular targets, and the binding mode to these targets. This review gives an overview of current knowledge on the ~2200 sequences, and more than 400 three-dimensional structures of snake toxins deposited in public repositories, as well as of molecular modeling studies of the interaction between these toxins and their molecular targets. We also describe how modern bioinformatics have been used to study the snake venom protein phospholipase A2, the small basic myotoxin Crotamine, and the three-finger peptide Mambalgin.

  5. Computational Studies of Snake Venom Toxins

    Directory of Open Access Journals (Sweden)

    Paola G. Ojeda

    2017-12-01

    Full Text Available Most snake venom toxins are proteins, and participate to envenomation through a diverse array of bioactivities, such as bleeding, inflammation, and pain, cytotoxic, cardiotoxic or neurotoxic effects. The venom of a single snake species contains hundreds of toxins, and the venoms of the 725 species of venomous snakes represent a large pool of potentially bioactive proteins. Despite considerable discovery efforts, most of the snake venom toxins are still uncharacterized. Modern bioinformatics tools have been recently developed to mine snake venoms, helping focus experimental research on the most potentially interesting toxins. Some computational techniques predict toxin molecular targets, and the binding mode to these targets. This review gives an overview of current knowledge on the ~2200 sequences, and more than 400 three-dimensional structures of snake toxins deposited in public repositories, as well as of molecular modeling studies of the interaction between these toxins and their molecular targets. We also describe how modern bioinformatics have been used to study the snake venom protein phospholipase A2, the small basic myotoxin Crotamine, and the three-finger peptide Mambalgin.

  6. Topological data analysis of Escherichia coli O157:H7 and non-O157 survival in soils

    Science.gov (United States)

    Shiga toxin-producing E. coli O157:H7 and non-O157 have been implicated in many foodborne illnesses caused by the consumption of contaminated fresh produce. However, data on their persistence in major fresh produce-growing soils are limited due to the complexity in datasets generated from different ...

  7. Pre-harvest management controls and intervention options for reducing escherichia coli O157:H7 shedding in cattle

    Science.gov (United States)

    Cattle can be naturally colonized with enterohemorrhagic E. coli (EHEC; also known as Shiga Toxin Producing E. coli, STEC) in their gastrointestinal tract. In order to further curtail these human illnesses and ensure a safe and wholesome food supply, research into preharvest E. coli O157:H7 and non...

  8. Environmental T4-Family Bacteriophages Evolve to Escape Abortive Infection via Multiple Routes in a Bacterial Host Employing "Altruistic Suicide" through Type III Toxin-Antitoxin Systems.

    Science.gov (United States)

    Chen, Bihe; Akusobi, Chidiebere; Fang, Xinzhe; Salmond, George P C

    2017-01-01

    Abortive infection is an anti-phage mechanism employed by a bacterium to initiate its own death upon phage infection. This reduces, or eliminates, production of viral progeny and protects clonal siblings in the bacterial population by an act akin to an "altruistic suicide." Abortive infection can be mediated by a Type III toxin-antitoxin system called ToxIN Pa consisting of an endoribonuclease toxin and RNA antitoxin. ToxIN Pa is a heterohexameric quaternary complex in which pseudoknotted RNA inhibits the toxicity of the toxin until infection by certain phages causes destabilization of ToxIN Pa , leading to bacteriostasis and, eventually, lethality. However, it is still unknown why only certain phages are able to activate ToxIN Pa . To try to address this issue we first introduced ToxIN Pa into the Gram-negative enterobacterium, Serratia sp. ATCC 39006 ( S 39006) and then isolated new environmental S 39006 phages that were scored for activation of ToxIN Pa and abortive infection capacity. We isolated three T4-like phages from a sewage treatment outflow point into the River Cam, each phage being isolated at least a year apart. These phages were susceptible to ToxIN Pa -mediated abortive infection but produced spontaneous "escape" mutants that were insensitive to ToxIN Pa . Analysis of these resistant mutants revealed three different routes of escaping ToxIN Pa , namely by mutating asiA (the product of which is a phage transcriptional co-activator); by mutating a conserved, yet functionally unknown, orf84 ; or by deleting a 6.5-10 kb region of the phage genome. Analysis of these evolved escape mutants may help uncover the nature of the corresponding phage product(s) involved in activation of ToxIN Pa .

  9. Clostridium difficile chimeric toxin receptor binding domain vaccine induced protection against different strains in active and passive challenge models.

    Science.gov (United States)

    Tian, Jing-Hui; Glenn, Gregory; Flyer, David; Zhou, Bin; Liu, Ye; Sullivan, Eddie; Wu, Hua; Cummings, James F; Elllingsworth, Larry; Smith, Gale

    2017-07-24

    Clostridium difficile is the number one cause of nosocomial antibiotic-associated diarrhea in developed countries. Historically, pathogenesis was attributed two homologous glucosylating toxins, toxin-A (TcdA) and toxin-B (TcdB). Over the past decade, however, highly virulent epidemic strains of C. difficile (B1/NAP1/027) have emerged and are linked to an increase in morbidity and mortality. Increased virulence is attributed to multiple factors including: increased production of A- and B-toxins; production of binary toxin (CDT); and the emergence of more toxic TcdB variants (TcdB (027) ). TcdB (027) is more cytotoxicity to cells; causes greater tissue damage and toxicity in animals; and is antigenically distinct from historical TcdB (TcdB (003) ). Broadly protective vaccines and therapeutic antibody strategies, therefore, may target TcdA, TcdB variants and CDT. To facilitate the generation of multivalent toxin-based C. difficile vaccines and therapeutic antibodies, we have generated fusion proteins constructed from the receptor binding domains (RBD) of TcdA, TcdB (003) , TcdB (027) and CDT. Herein, we describe the development of a trivalent toxin (T-toxin) vaccine (CDTb/TcdB (003) /TcdA) and quadravalent toxin (Q-toxin) vaccine (CDTb/TcB (003) /TcdA/TcdB (027) ) fusion proteins that retain the protective toxin neutralizing epitopes. Active immunization of mice or hamsters with T-toxin or Q-toxin fusion protein vaccines elicited the generation of toxin neutralizing antibodies to each of the toxins. Hamsters immunized with the Q-toxin vaccine were broadly protected against spore challenge with historical C. difficile 630 (toxinotype 0/ribotype 003) and epidemic NAP1 (toxinotype III/ribotype 027) strains. Fully human polyclonal antitoxin IgG was produced by immunization of transgenic bovine with these fusion proteins. In passive transfer studies, mice were protected against lethal toxin challenge. Hamsters treated with human antitoxin IgG were completely protected when

  10. Nutrients and toxin producing phytoplankton control algal blooms - a spatio-temporal study in a noisy environment.

    Science.gov (United States)

    Sarkar, Ram Rup; Malchow, Horst

    2005-12-01

    A phytoplankton-zooplankton prey-predator model has been investigated for temporal, spatial and spatio-temporal dissipative pattern formation in a deterministic and noisy environment, respectively. The overall carrying capacity for the phytoplankton population depends on the nutrient level. The role of nutrient concentrations and toxin producing phytoplankton for controlling the algal blooms has been discussed. The local analysis yields a number of stationary and/or oscillatory regimes and their combinations. Correspondingly interesting is the spatio-temporal behaviour, modelled by stochastic reaction-diffusion equations. The present study also reveals the fact that the rate of toxin production by toxin producing phytoplankton (TPP) plays an important role for controlling oscillations in the plankton system. We also observe that different mortality functions of zooplankton due to TPP have significant influence in controlling oscillations, coexistence, survival or extinction of the zoo-plankton population. External noise can enhance the survival and spread of zooplankton that would go extinct in the deterministic system due to a high rate of toxin production.

  11. 9 CFR 121.4 - Overlap select agents and toxins.

    Science.gov (United States)

    2010-01-01

    ... in paragraph (b) of this section that have been genetically modified. (d) Overlap select agents or... OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS POSSESSION, USE... elements, recombinant nucleic acids, and recombinant organisms: (1) Nucleic acids that can produce...

  12. Brown spider dermonecrotic toxin directly induces nephrotoxicity

    International Nuclear Information System (INIS)

    Chaim, Olga Meiri; Sade, Youssef Bacila; Bertoni da Silveira, Rafael; Toma, Leny; Kalapothakis, Evanguedes; Chavez-Olortegui, Carlos; Mangili, Oldemir Carlos; Gremski, Waldemiro; Dietrich, Carl Peter von; Nader, Helena B.; Sanches Veiga, Silvio

    2006-01-01

    Brown spider (Loxosceles genus) venom can induce dermonecrotic lesions at the bite site and systemic manifestations including fever, vomiting, convulsions, disseminated intravascular coagulation, hemolytic anemia and acute renal failure. The venom is composed of a mixture of proteins with several molecules biochemically and biologically well characterized. The mechanism by which the venom induces renal damage is unknown. By using mice exposed to Loxosceles intermedia recombinant dermonecrotic toxin (LiRecDT), we showed direct induction of renal injuries. Microscopic analysis of renal biopsies from dermonecrotic toxin-treated mice showed histological alterations including glomerular edema and tubular necrosis. Hyalinization of tubules with deposition of proteinaceous material in the tubule lumen, tubule epithelial cell vacuoles, tubular edema and epithelial cell lysis was also observed. Leukocytic infiltration was neither observed in the glomerulus nor the tubules. Renal vessels showed no sign of inflammatory response. Additionally, biochemical analyses showed such toxin-induced changes in renal function as urine alkalinization, hematuria and azotemia with elevation of blood urea nitrogen levels. Immunofluorescence with dermonecrotic toxin antibodies and confocal microscopy analysis showed deposition and direct binding of this toxin to renal intrinsic structures. By immunoblotting with a hyperimmune dermonecrotic toxin antiserum on renal lysates from toxin-treated mice, we detected a positive signal at the region of 33-35 kDa, which strengthens the idea that renal failure is directly induced by dermonecrotic toxin. Immunofluorescence reaction with dermonecrotic toxin antibodies revealed deposition and binding of this toxin directly in MDCK epithelial cells in culture. Similarly, dermonecrotic toxin treatment caused morphological alterations of MDCK cells including cytoplasmic vacuoles, blebs, evoked impaired spreading and detached cells from each other and from

  13. The Agricultural Antibiotic Carbadox Induces Phage-mediated Gene Transfer in Salmonella

    Directory of Open Access Journals (Sweden)

    Bradley L. Bearson

    2014-02-01

    Full Text Available Antibiotics are used for disease therapeutic or preventative effects in humans and animals, as well as for enhanced feed conversion efficiency in livestock. Antibiotics can also cause undesirable effects in microbial populations, including selection for antibiotic resistance, enhanced pathogen invasion, and stimulation of horizontal gene transfer. Carbadox is a veterinary antibiotic used in the U.S. during the starter phase of swine production for improved feed efficiency and control of swine dysentery and bacterial swine enteritis. Carbadox has been shown in vitro to induce phage-encoded Shiga toxin in Shiga toxin-producing Escherichia coli and a phage-like element transferring antibiotic resistance genes in Brachyspira hyodysenteriae, but the effect of carbadox on prophages in other bacteria is unknown. This study examined carbadox exposure on prophage induction and genetic transfer in Salmonella enterica serovar Typhimurium, a human foodborne pathogen that frequently colonizes swine without causing disease. S. Typhimurium LT2 exposed to carbadox induced prophage production, resulting in bacterial cell lysis and release of virions that were visible by electron microscopy. Carbadox induction of phage-mediated gene transfer was confirmed by monitoring the transduction of a sodCIII::neo cassette in the Fels-1 prophage from LT2 to a recipient Salmonella strain. Furthermore, carbadox frequently induced generalized transducing phages in multidrug-resistant phage type DT104 and DT120 isolates, resulting in the transfer of chromosomal and plasmid DNA that included antibiotic resistance genes. Our research indicates that exposure of Salmonella to carbadox induces prophages that can transfer virulence and antibiotic resistance genes to susceptible bacterial hosts. Carbadox-induced, phage-mediated gene transfer could serve as a contributing factor in bacterial evolution during animal production, with prophages being a reservoir for bacterial fitness

  14. Cellular Entry of Clostridium perfringens Iota-Toxin and Clostridium botulinum C2 Toxin

    Directory of Open Access Journals (Sweden)

    Masaya Takehara

    2017-08-01

    Full Text Available Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin are composed of two non-linked proteins, one being the enzymatic component and the other being the binding/translocation component. These latter components recognize specific receptors and oligomerize in plasma membrane lipid-rafts, mediating the uptake of the enzymatic component into the cytosol. Enzymatic components induce actin cytoskeleton disorganization through the ADP-ribosylation of actin and are responsible for cell rounding and death. This review focuses upon the recent advances in cellular internalization of clostridial binary toxins.

  15. EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin.

    Science.gov (United States)

    Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Azarnia Tehran, Domenico; Montecucco, Cesare; Barth, Holger

    2016-04-01

    The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins.

  16. EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin

    Science.gov (United States)

    Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R.; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Tehran, Domenico Azarnia; Montecucco, Cesare; Barth, Holger

    2016-01-01

    The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins. PMID:27043629

  17. Cross-talk between light and glucose regulation controls toxin production and morphogenesis in Aspergillus nidulans

    International Nuclear Information System (INIS)

    Atoui, A.; Larey, C.; Thokala, R.; Calvo, A.M.; Kastner, C.; Fischer, R.; Etxebeste, O; Espeso, E.A.

    2010-01-01

    Light is a major environmental stimulus that has a broad effect on organisms, triggering a cellular response that results in an optimal adaptation enhancing fitness and survival. In fungi, light affects growth, and causes diverse morphological changes such as those leading to reproduction. Light can also affect fungal metabolism, including the biosynthesis of natural products. In this study we show that in Aspergillus nidulans the effect of light on the production of the sterigmatocystin (ST) toxin depends on the glucose concentration. In cultures grown with 1% glucose and exposed to light, ST production was lower than when grown in the dark. This lower ST production coincided with an elevated rate of cellular damage with partial loss of nuclear integrity and vacuolated cytoplasm. However, in cultures grown with 2% glucose these effects were reversed and light enhanced ST production. Glucose abundance also affected the light-dependent subcellular localization of the VeA (velvet) protein, a key regulator necessary for normal light-dependent morphogenesis and secondary metabolism in Aspergilli and other fungal gen- era. The role of other VeA-associated proteins, particularly the blue-light-sensing proteins LreA and LreB (WC-1 and WC-2 orthologs), on conidiation could also be modified by the abundance of glucose. We also show that LreA and LreB, as well as the phytochrome FphA, modulate not only the synthesis of sterigmat- ocystin, but also the production of the antibiotic penicillin. (author)

  18. Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

    International Nuclear Information System (INIS)

    Rose, S.A.; Bailey, N.E.; Stringer, M.F.; Modi, N.K.; Tranter, H.S.; Hambleton, P.

    1989-01-01

    The effects of irradiation of Clostridium botulinum neutotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author)

  19. Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

    Energy Technology Data Exchange (ETDEWEB)

    Rose, S A; Bailey, N E; Stringer, M F [Campden Food and Drink Research Association, Chipping Campden (UK); Modi, N K; Tranter, H S [Porton International plc., London (UK); Hambleton, P [Centre for Applied Microbiological Research, Porton Down (UK)

    1988-10-01

    The effects of irradiation of Clostridium botulinum neurotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author).

  20. Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

    International Nuclear Information System (INIS)

    Rose, S.A.; Bailey, N.E.; Stringer, M.F.; Modi, N.K.; Tranter, H.S.; Hambleton, P.

    1988-01-01

    The effects of irradiation of Clostridium botulinum neurotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author)