Association of Escherichia coli O157:H7 with Preharvest Leaf Lettuce upon Exposure to Contaminated Irrigation Water

2002 ◽  
Vol 65 (1) ◽  
pp. 18-25 ◽  
Author(s):  
MARIAN R. WACHTEL ◽  
LINDA C. WHITEHAND ◽  
ROBERT E. MANDRELL
Keyword(s):  
Escherichia Coli ◽  
Irrigation Water ◽  
Enterohemorrhagic Escherichia Coli ◽  
Laser Scanning Microscopy ◽  
Model Systems ◽  
Plant Roots ◽  
Confocal Laser ◽  
Food Handler ◽  
Hydroponic System ◽  
Contaminated Irrigation Water

Recent foodborne outbreaks have linked infection by enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 to the consumption of contaminated lettuce. Contamination via food handler error and on-the-farm contamination are thought to be responsible for several outbreaks. Though recent studies have examined the application of EHEC to store-bought lettuce, little is known about the attachment of EHEC to growing plants. We investigated the association of lettuce seedlings with EHEC O157:H7 strains implicated in lettuce or fruit outbreaks using hydroponic and soil model systems. EHEC strains that express the green fluorescent protein were observed by stereomicroscopy and confocal laser scanning microscopy to determine adherence patterns on growing lettuce seedlings. Bacteria adhered preferentially to plant roots in both model systems and to seed coats in the hydroponic system. Two of five nonpathogenic E. coli strains showed decreased adherence to seedling roots in the hydroponic system. EHEC was associated with plants in as few as 3 days in soil, and contamination levels were dose-dependent. EHEC levels associated with young plants inoculated with a low dose suggested that the bacteria had multiplied. These data suggest that preharvest crop contamination via contaminated irrigation water can occur through plant roots.

scholarly journals Two Enteropathogenic Escherichia coli Type III Secreted Proteins, EspA and EspB, Are Virulence Factors

1998 ◽  
Vol 188 (10) ◽  
pp. 1907-1916 ◽  
Author(s):  
Akio Abe ◽  
Ursula Heczko ◽  
Richard G. Hegele ◽  
B. Brett Finlay
Keyword(s):  
Escherichia Coli ◽  
Virulence Factors ◽  
Enterohemorrhagic Escherichia Coli ◽  
Laser Scanning Microscopy ◽  
Secreted Proteins ◽  
Confocal Laser ◽  
Infection Model ◽  
Enteropathogenic Escherichia Coli ◽  
Type Iii

Enteropathogenic Escherichia coli (EPEC) belongs to a family of related bacterial pathogens, including enterohemorrhagic Escherichia coli (EHEC) O157:H7 and other human and animal diarrheagenic pathogens that form attaching and effacing (A/E) lesions on host epithelial surfaces. Bacterial secreted Esp proteins and a type III secretion system are conserved among these pathogens and trigger host cell signal transduction pathways and cytoskeletal rearrangements, and mediate intimate bacterial adherence to epithelial cell surfaces in vitro. However, their role in pathogenesis is still unclear. To investigate the role of Esp proteins in disease, mutations in espA and espB were constructed in rabbit EPEC serotype O103 and infection characteristics were compared to that of the wild-type strain using histology, scanning and transmission electron microscopy, and confocal laser scanning microscopy in a weaned rabbit infection model. The virulence of EspA and EspB mutant strains was severely attenuated. Additionally, neither mutant strain formed A/E lesions, nor did either one cause cytoskeletal actin rearrangements beneath the attached bacteria in the rabbit intestine. Collectively, this study shows for the first time that the type III secreted proteins EspA and EspB are needed to form A/E lesions in vivo and are indeed virulence factors. It also confirms the role of A/E lesions in disease processes.

Persistence of Enterohemorrhagic Escherichia coli O157:H7 in Soil and on Leaf Lettuce and Parsley Grown in Fields Treated with Contaminated Manure Composts or Irrigation Water

2004 ◽  
Vol 67 (7) ◽  
pp. 1365-1370 ◽  
Author(s):  
MAHBUB ISLAM ◽  
MICHAEL P. DOYLE ◽  
SHARAD C. PHATAK ◽  
PATRICIA MILLNER ◽  
XIUPING JIANG
Keyword(s):  
Escherichia Coli ◽  
Irrigation Water ◽  
Poultry Manure ◽  
Dairy Manure ◽  
Enterohemorrhagic Escherichia Coli ◽  
Avirulent Strain ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Manure Compost ◽  
Contaminated Irrigation Water

Outbreaks of enterohemorrhagic Escherichia coli O157:H7 infections associated with lettuce and other leaf crops have occurred with increasing frequency in recent years. Contaminated manure and polluted irrigation water are probable vehicles for the pathogen in many outbreaks. In this study, the occurrence and persistence of E. coli O157:H7 in soil fertilized with contaminated poultry or bovine manure composts or treated with contaminated irrigation water and on lettuce and parsley grown on these soils under natural environmental conditions was determined. Twenty-five plots, each 1.8 by 4.6 m, were used for each crop, with five treatments (one without compost, three with each of the three composts, and one without compost but treated with contaminated water) and five replication plots for each treatment. Three different types of compost, PM-5 (poultry manure compost), 338 (dairy manure compost), and NVIRO-4 (alkaline-stabilized dairy manure compost), and irrigation water were inoculated with an avirulent strain of E. coli O157:H7. Pathogen concentrations were 107 CFU/g of compost and 105 CFU/ml of water. Contaminated compost was applied to soil in the field as a strip at 4.5 metric tons per hectare on the day before lettuce and parsley seedlings were transplanted in late October 2002. Contaminated irrigation water was applied only once on the plants as a treatment in five plots for each crop at the rate of 2 liters per plot 3 weeks after the seedlings were transplanted. E. coli O157:H7 persisted for 154 to 217 days in soils amended with contaminated composts and was detected on lettuce and parsley for up to 77 and 177 days, respectively, after seedlings were planted. Very little difference was observed in E. coli O157:H7 persistence based on compost type alone. E. coli O157:H7 persisted longer (by >60 days) in soil covered with parsley plants than in soil from lettuce plots, which were bare after lettuce was harvested. In all cases, E. coli O157:H7 in soil, regardless of source or crop type, persisted for >5 months after application of contaminated compost or irrigation water.

Surface and Internalized Escherichia coli O157:H7 on Field-Grown Spinach and Lettuce Treated with Spray-Contaminated Irrigation Water

2010 ◽  
Vol 73 (6) ◽  
pp. 1023-1029 ◽  
Author(s):  
MARILYN C. ERICKSON ◽  
CATHY C. WEBB ◽  
JUAN CARLOS DIAZ-PEREZ ◽  
SHARAD C. PHATAK ◽  
JOHN J. SILVOY ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Irrigation Water ◽  
Enrichment Culture ◽  
Field Studies ◽  
Plate Count ◽  
Escherichia Coli O157 ◽  
Adaxial Side ◽  
Abaxial Side ◽  
E Coli ◽  
Contaminated Irrigation Water

Numerous field studies have revealed that irrigation water can contaminate the surface of plants; however, the occurrence of pathogen internalization is unclear. This study was conducted to determine the sites of Escherichia coli O157:H7 contamination and its survival when the bacteria were applied through spray irrigation water to either field-grown spinach or lettuce. To differentiate internalized and surface populations, leaves were treated with a surface disinfectant wash before the tissue was ground for analysis of E. coli O157:H7 by direct plate count or enrichment culture. Irrigation water containing E. coli O157:H7 at 102, 104, or 106 CFU/ml was applied to spinach 48 and 69 days after transplantation of seedlings into fields. E. coli O157:H7 was initially detected after application on the surface of plants dosed at 104 CFU/ml (4 of 20 samples) and both on the surface (17 of 20 samples) and internally (5 of 20 samples) of plants dosed at 106 CFU/ml. Seven days postspraying, all spinach leaves tested negative for surface or internal contamination. In a subsequent study, irrigation water containing E. coli O157:H7 at 108 CFU/ml was sprayed onto either the abaxial (lower) or adaxial (upper) side of leaves of field-grown lettuce under sunny or shaded conditions. E. coli O157:H7 was detectable on the leaf surface 27 days postspraying, but survival was higher on leaves sprayed on the abaxial side than on leaves sprayed on the adaxial side. Internalization of E. coli O157:H7 into lettuce leaves also occurred with greater persistence in leaves sprayed on the abaxial side (up to 14 days) than in leaves sprayed on the adaxial side (2 days).

Infrequent Internalization of Escherichia coli O157:H7 into Field-Grown Leafy Greens

2010 ◽  
Vol 73 (3) ◽  
pp. 500-506 ◽  
Author(s):  
MARILYN C. ERICKSON ◽  
CATHY C. WEBB ◽  
JUAN CARLOS DIAZ-PEREZ ◽  
SHARAD C. PHATAK ◽  
JOHN J. SILVOY ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Irrigation Water ◽  
Field Studies ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Leafy Greens ◽  
Contaminated Irrigation Water ◽  
Green Fluorescent ◽  
Field Plots ◽  
Spinach Leaves

Several sources of contamination of fresh produce by Escherichia coli O157:H7 (O157) have been identified and include contaminated irrigation water and improperly composted animal waste; however, field studies evaluating the potential for internalization of O157 into leafy greens from these sources have not been conducted. Irrigation water inoculated with green fluorescent plasmid–labeled Shiga toxin–negative strains (50 ml of 102, 104, or 106 CFU of O157 per ml) was applied to soil at the base of spinach plants of different maturities in one field trial. In a second trial, contaminated compost (1.8 kg of 103 or 105 CFU of O157 per g) was applied to field plots (0.25 by 3.0 m) prior to transplantation of spinach, lettuce, or parsley plants. E. coli O157:H7 persisted in the soil up to harvest (day 76 posttransplantation) following application of contaminated irrigation water; however, internalized O157 was not detected in any spinach leaves or in roots exposed to O157 during the early or late growing season. Internalized O157 was detected in root samples collected 7 days after plants were contaminated in mid-season, with 5 of 30 samples testing positive for O157 by enrichment; however, O157 was not detected by enrichment in surface-disinfected roots on days 14 or 22. Roots and leaves from transplanted spinach, lettuce, and parsley did not internalize O157 for up to 50 days in the second trial. These results indicate that internalization of O157 via plant roots in the field is rare and when it does occur, O157 does not persist 7 days later.

scholarly journals The Chromosomal Toxin Gene yafQ Is a Determinant of Multidrug Tolerance for Escherichia coli Growing in a Biofilm

2009 ◽  
Vol 53 (6) ◽  
pp. 2253-2258 ◽  
Author(s):  
Joe J. Harrison ◽  
William D. Wade ◽  
Sarah Akierman ◽  
Caterina Vacchi-Suzzi ◽  
Carol A. Stremick ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Cell Survival ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Cell Counting ◽  
Confocal Laser ◽  
Toxin Gene ◽  
Persister Cells ◽  
E Coli

ABSTRACT Escherichia coli is refractory to elevated doses of antibiotics when it is growing in a biofilm, and this is potentially due to high numbers of multidrug-tolerant persister cells in the surface-adherent population. Previously, the chromosomal toxin-antitoxin loci hipBA and relBE have been linked to the frequency at which persister cells occur in E. coli populations. In the present study, we focused on the dinJ-yafQ-encoded toxin-antitoxin system and hypothesized that deletion of the toxin gene yafQ might influence cell survival in antibiotic-exposed biofilms. By using confocal laser scanning microscopy and viable cell counting, it was determined that a ΔyafQ mutant produced biofilms with a structure and a cell density equivalent to those of the parental strain. In-depth susceptibility testing identified that relative to wild-type E. coli, the ΔyafQ strain had up to a ∼2,400-fold decrease in cell survival after the biofilms were exposed to bactericidal concentrations of cefazolin or tobramycin. Corresponding to these data, controlled overexpression of yafQ from a high-copy-number plasmid resulted in up to a ∼10,000-fold increase in the number of biofilm cells surviving exposure to these bactericidal drugs. In contrast, neither the inactivation nor the overexpression of yafQ affected the tolerance of biofilms to doxycycline or rifampin (rifampicin). Furthermore, deletion of yafQ did not affect the tolerance of stationary-phase planktonic cells to any of the antibacterials tested. These results suggest that yafQ mediates the tolerance of E. coli biofilms to multiple but specific antibiotics; moreover, our data imply that this cellular pathway for persistence is likely different from that of multidrug-tolerant cells in stationary-phase planktonic cell cultures.

scholarly journals Antibody response to the 45 kDa Candida albicans antigen in an animal model and potential role of the antigen in adherence

2008 ◽  
Vol 57 (12) ◽  
pp. 1466-1472 ◽  
Author(s):  
Helena Bujdáková ◽  
Ema Paulovičová ◽  
Silvia Borecká-Melkusová ◽  
Juraj Gašperík ◽  
Soňa Kucharíková ◽  
...  
Keyword(s):  
Animal Model ◽  
Candida Albicans ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Vaccine Development ◽  
Laser Scanning Microscopy ◽  
Model Systems ◽  
Confocal Laser

The Candida antigen CR3-RP (complement receptor 3-related protein) is supposed to be a ‘mimicry’ protein because of its ability to bind antibody directed against the α subunit of the mammalian CR3 (CD11b/CD18). This study aimed to (i) investigate the specific humoral isotypic response to immunization with CR3-RP in vivo in a rabbit animal model, and (ii) determine the role of CR3-RP in the adherence of Candida albicans in vitro using the model systems of buccal epithelial cells (BECs) and biofilm formation. The synthetic C. albicans peptide DINGGGATLPQ corresponding to 11 amino-acids of the CR3-RP sequence DINGGGATLPQALXQITGVIT, determined by N-terminal sequencing, was used for immunization of rabbits to obtain polyclonal anti-CR3-PR serum and for subsequent characterization of the humoral isotypic response of rabbits. A significant increase of IgG, IgA and IgM anti-CR3-RP specific antibodies was observed after the third (P<0.01) and the fourth (P<0.001) immunization doses. The elevation of IgA levels suggested peptide immunomodulation of the IgA1 subclass, presumably in coincidence with Candida epithelial adherence. Blocking CR3-RP with polyclonal anti-CR3-RP serum reduced the ability of Candida to adhere to BECs, in comparison with the control, by up to 35 % (P<0.001), and reduced biofilm formation by 28 % (P<0.001), including changes in biofilm thickness and integrity detected by confocal laser scanning microscopy. These properties of CR3-RP suggest that it has potential for future vaccine development.

scholarly journals Food Microstructure and Fat Content Affect Growth Morphology, Growth Kinetics, and Preferred Phase for Cell Growth ofListeria monocytogenesin Fish-Based Model Systems

2019 ◽  
Vol 85 (16) ◽  
Author(s):  
Davy Verheyen ◽  
Xiang Ming Xu ◽  
Marlies Govaert ◽  
Maria Baka ◽  
Torstein Skåra ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Microbial Growth ◽  
Growth Dynamics ◽  
Growth Behavior ◽  
Laser Scanning Microscopy ◽  
Model Systems ◽  
Confocal Laser ◽  
Growth Morphology ◽  
Content Type ◽  
Food Model

ABSTRACTFood microstructure significantly affects microbial growth dynamics, but knowledge concerning the exact influencing mechanisms at a microscopic scale is limited. The food microstructural influence onListeria monocytogenes(green fluorescent protein strain) growth at 10°C in fish-based food model systems was investigated by confocal laser scanning microscopy. The model systems had different microstructures, i.e., liquid, xanthan (high-viscosity liquid), aqueous gel, and emulsion and gelled emulsion systems varying in fat content. Bacteria grew as single cells, small aggregates, and microcolonies of different sizes (based on colony radii [size I, 1.5 to 5.0 μm; size II, 5.0 to 10.0 μm; size III, 10.0 to 15.0 μm; and size IV, ≥15 μm]). In the liquid, small aggregates and size I microcolonies were predominantly present, while size II and III microcolonies were predominant in the xanthan and aqueous gel. Cells in the emulsions and gelled emulsions grew in the aqueous phase and on the fat-water interface. A microbial adhesion to solvent assay demonstrated limited bacterial nonpolar solvent affinities, implying that this behavior was probably not caused by cell surface hydrophobicity. In systems containing 1 and 5% fat, the largest cell volume was mainly represented by size I and II microcolonies, while at 10 and 20% fat a few size IV microcolonies comprised nearly the total cell volume. Microscopic results (concerning, e.g., growth morphology, microcolony size, intercolony distances, and the preferred phase for growth) were related to previously obtained macroscopic growth dynamics in the model systems for anL. monocytogenesstrain cocktail, leading to more substantiated explanations for the influence of food microstructural aspects on lag phase duration and growth rate.IMPORTANCEListeria monocytogenesis one of the most hazardous foodborne pathogens due to the high fatality rate of the disease (i.e., listeriosis). In this study, the growth behavior ofL. monocytogeneswas investigated at a microscopic scale in food model systems that mimic processed fish products (e.g., fish paté and fish soup), and the results were related to macroscopic growth parameters. Many studies have previously focused on the food microstructural influence on microbial growth. The novelty of this work lies in (i) the microscopic investigation of products with a complex composition and/or structure using confocal laser scanning microscopy and (ii) the direct link to the macroscopic level. Growth behavior (i.e., concerning bacterial growth morphology and preferred phase for growth) was more complex than assumed in common macroscopic studies. Consequently, the effectiveness of industrial antimicrobial food preservation technologies (e.g., thermal processing) might be overestimated for certain products, which may have critical food safety implications.

scholarly journals Neutral Red as a Probe for Confocal Laser Scanning Microscopy Studies of Plant Roots

2006 ◽  
Vol 97 (6) ◽  
pp. 1127-1138 ◽  
Author(s):  
JOSEPH G. DUBROVSKY ◽  
MARTIN GUTTENBERGER ◽  
ANDRES SARALEGUI ◽  
SELENE NAPSUCIALY-MENDIVIL ◽  
BORIS VOIGT ◽  
...  
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Neutral Red ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Plant Roots ◽  
Confocal Laser

scholarly journals Inhibition of Escherichia coli Biofilm Formation by Self-Assembled Monolayers of Functional Alkanethiols on Gold

2007 ◽  
Vol 73 (13) ◽  
pp. 4300-4307 ◽  
Author(s):  
Shuyu Hou ◽  
Erik A. Burton ◽  
Karen A. Simon ◽  
Dustin Blodgett ◽  
Yan-Yeung Luk ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Amide Bond ◽  
Self Assembled Monolayers ◽  
Confocal Laser ◽  
E Coli ◽  
Assembled Monolayers ◽  
Self Assembled

ABSTRACT Bacterial biofilms cause serious problems, such as antibiotic resistance and medical device-related infections. To further understand bacterium-surface interactions and to develop efficient control strategies, self-assembled monolayers (SAMs) of alkanethiols presenting different functional groups on gold films were analyzed to determine their resistance to biofilm formation. Escherichia coli was labeled with green florescence protein, and its biofilm formation on SAM-modified surfaces was monitored by confocal laser scanning microscopy. The three-dimensional structures of biofilms were analyzed with the COMSTAT software to obtain information about biofilm thickness and surface coverage. SAMs presenting methyl, l-gulonamide (a sugar alcohol tethered with an amide bond), and tri(ethylene glycol) (TEG) groups were tested. Among these, the TEG-terminated SAM was the most resistant to E. coli biofilm formation; e.g., it repressed biofilm formation by E. coli DH5α by 99.5% ± 0.1% for 1 day compared to the biofilm formation on a bare gold surface. When surfaces were patterned with regions consisting of methyl-terminated SAMs surrounded by TEG-terminated SAMs, E. coli formed biofilms only on methyl-terminated patterns. Addition of TEG as a free molecule to growth medium at concentrations of 0.1 and 1.0% also inhibited biofilm formation, while TEG at concentrations up to 1.5% did not have any noticeable effects on cell growth. The results of this study suggest that the reduction in biofilm formation on surfaces modified with TEG-terminated SAMs is a result of multiple factors, including the solvent structure at the interface, the chemorepellent nature of TEG, and the inhibitory effect of TEG on cell motility.

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