Affinity isolation and I-DIRT mass spectrometric analysis of the Escherichia coli O157:H7 Sakai RNA polymerase complex.

Science.gov (United States)

Lee, David J; Busby, Stephen J W; Westblade, Lars F; Chait, Brian T

2008-02-01

Bacteria contain a single multisubunit RNA polymerase that is responsible for the synthesis of all RNA. Previous studies of the Escherichia coli K-12 laboratory strain identified a group of effector proteins that interact directly with RNA polymerase to modulate the efficiency of transcription initiation, elongation, or termination. Here we used a rapid affinity isolation technique to isolate RNA polymerase from the pathogenic Escherichia coli strain O157:H7 Sakai. We analyzed the RNA polymerase enzyme complex using mass spectrometry and identified associated proteins. Although E. coli O157:H7 Sakai contains more than 1,600 genes not present in the K-12 strain, many of which are predicted to be involved in transcription regulation, all of the identified proteins in this study were encoded on the "core" E. coli genome.

Variation in Resistance of Natural Isolates of Escherichia coli O157 to High Hydrostatic Pressure, Mild Heat, and Other Stresses

OpenAIRE

Benito, Amparo; Ventoura, Georgia; Casadei, Maria; Robinson, Tobin; Mackey, Bernard

1999-01-01

Strains of Escherichia coli O157 isolated from patients with clinical cases of food-borne illness and other sources exhibited wide differences in resistance to high hydrostatic pressure. The most pressure-resistant strains were also more resistant to mild heat than other strains. Strain C9490, a representative pressure-resistant strain, was also more resistant to acid, oxidative, and osmotic stresses than the pressure-sensitive strain NCTC 12079. Most of these differences in resistance were o...

Prevalence and antibiogram of Escherichia coli O157 isolated from ...

African Journals Online (AJOL)

E. coli O157 is an important serotype that caused many food borne outbreaks worldwide in the past decades. This study was carried out to estimate the prevalence and determine the antimicrobial susceptibility of E. coli O157 isolated from bovine carcasses and cecal contents at one abattoir in Jimma. A total of 300 samples ...

Aislamiento, caracterización y subtipificación de cepas de Escherichia coli O157:H7 a partir de productos cárnicos y leche Isolation, characterization and typing of Escherichia coli O157:H7 strains from beef products and milk

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M. L. Roldán

2007-06-01

Full Text Available Escherichia coli productor de toxina Shiga (Stx (STEC O157:H7 es un patógeno asociado a enfermedades transmitidas por alimentos, fundamentalmente de origen animal. Se investigó la presencia de E. coli O157 en 250 muestras de carne picada y hamburguesas obtenidas de comercios de las ciudades de Santa Fe y Santo Tomé (Pcia. de Santa Fe y en 150 muestras de leche provenientes de tanques de enfriado de tambos de la región, utilizando enriquecimiento selectivo y separación inmunomagnética. A partir de 3 muestras de carne (1,2% se aislaron cepas E. coli O157:H7 stx2, eae, y ehxA positivas, que pudieron ser diferenciadas mediante electroforesis de campo pulsado, fagotipificación y genotipificación de stx. No se aislaron cepas STEC O157:H7 a partir de las muestras de leche. Estos hallazgos confirman la participación de los alimentos de origen animal en la epidemiología de las enfermedades producidas por E. coli O157:H7.Shiga toxin (Stx-producing Escherichia coli (STEC is an emergent pathogen associated with foodborne diseases, especially foodstuffs of animal origin. A total of 250 beef samples (ground beef and hamburgers obtained from retail outlets in Santa Fe and Santo Tomé cities, and 150 milk samples from bulk tank milk from dairy barns of the region were analyzed by selective enrichment and immunomagnetic separation. Escherichia coli O157:H7 stx2, eae and ehxA positive strains were isolated from three (1.2% beef samples. The strains could be differentiated by pulsed-field gel electrophoresis, phagetyping and genotyping of stx. The milk samples were negative for STEC O157. These findings confirm the role of food of animal origin in the epidemiology of E. coli O157:H7 - associated diseases.

Irradiation effect of escherichia coli O157 : H7 in meats

International Nuclear Information System (INIS)

Ito, Hitoshi; Harsojo

1998-01-01

Escherichia coli O157 : H7 is a rapidly emerging food-born pathogen which has been linked to outbreaks of hemorrhagic diarrhea in Japan, USA or many European countries. From this study, two strains of E. coli O157 : H7 were isolated from beef and chicken meats in each one sample of 4 replicates. Some of the biochemical characteristics of these isolates were different from type strain IID959. These isolates could grow quickly at 10degC on cultivation of nutrient agar. D 10 value of these isolates were obtained to be 0.06kGy in 0.067M phosphate buffer suspension which were highly sensitive than D 10 value of 0.12kGy on type strain IID959. On the irradiation effect of type strain IID959 in ground beef, D 10 value was obtained as 0.26kGy at fresh condition and 0.46kGy at frozen condition, respectively. From these results, necessary dose for elimination of E. coli O157 : H7 was decided as 1.5kGy for fresh meats and 3kGy for frozen meats. (author)

Affinity Isolation and I-DIRT Mass Spectrometric Analysis of the Escherichia coli O157:H7 Sakai RNA Polymerase Complex▿

Science.gov (United States)

Lee, David J.; Busby, Stephen J. W.; Westblade, Lars F.; Chait, Brian T.

2008-01-01

Bacteria contain a single multisubunit RNA polymerase that is responsible for the synthesis of all RNA. Previous studies of the Escherichia coli K-12 laboratory strain identified a group of effector proteins that interact directly with RNA polymerase to modulate the efficiency of transcription initiation, elongation, or termination. Here we used a rapid affinity isolation technique to isolate RNA polymerase from the pathogenic Escherichia coli strain O157:H7 Sakai. We analyzed the RNA polymerase enzyme complex using mass spectrometry and identified associated proteins. Although E. coli O157:H7 Sakai contains more than 1,600 genes not present in the K-12 strain, many of which are predicted to be involved in transcription regulation, all of the identified proteins in this study were encoded on the “core” E. coli genome. PMID:18083804

Genomic Variability of O Islands Encoding Tellurite Resistance in Enterohemorrhagic Escherichia coli O157:H7 Isolates

OpenAIRE

Taylor, Diane E.; Rooker, Michelle; Keelan, Monika; Ng, Lai-King; Martin, Irene; Perna, Nicole T.; Burland, N. T. Valerie; Blattner, Fredrick R.

2002-01-01

Strains of Escherichia coli causing enterohemorrhagic colitis belonging to the O157:H7 lineage are reported to be highly related. Fifteen strains of E. coli O157:H7 and 1 strain of E. coli O46:H− (nonflagellated) were examined for the presence of potassium tellurite resistance (Ter). Ter genes comprising terABCDEF were shown previously to be part of a pathogenicity island also containing integrase, phage, and urease genes. PCR analysis, both conventional and light cycler based, demonstrated t...

[Construction and characterization of enterohemorrhagic Escherichia coli O157:H7 ppk- deleted strain].

Science.gov (United States)

Han, Peng; Sun, Qi; Zhao, Suhui; Zhang, Qiwei; Wan, Chengsong

2014-06-01

To construct enterohemorrhagic Escherichia coli (EHEC) O157: H7 ppk gene deletion strains and study its biological characteristics. The gene fragment of kanamycin resistance was amplified using a pair of homologous arm primers whose 5' and 3' ends were homologous with ppk gene and kanamycin resistance gene, respectively. EHEC O157: H7 EDL933w competent strains were prepared and transformed via electroporation with the amplification products. The ppk gene was replaced by kanamycin resistance gene using pKD46-mediated Red recombination system. The recombinant strain was confirmed by PCR and sequencing, and its morphology, growth ability and adhesion were assessed using Gram staining, OD600 value and Giemsa staining. We established a ppk-deleted EHEC O157:H7 EDL933w strain with kanamycin resistance and compared the biological characteristics of the wild-type and mutant strains, which may facilitate further study of the regulatory mechanism of ppk gene.

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)

Overexpressed Proteins in Hypervirulent Clade 8 and Clade 6 Strains of Escherichia coli O157:H7 Compared to E. coli O157:H7 EDL933 Clade 3 Strain.

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Natalia Amigo

Full Text Available Escherichia coli O157:H7 is responsible for severe diarrhea and hemolytic uremic syndrome (HUS, and predominantly affects children under 5 years. The major virulence traits are Shiga toxins, necessary to develop HUS and the Type III Secretion System (T3SS through which bacteria translocate effector proteins directly into the host cell. By SNPs typing, E. coli O157:H7 was separated into nine different clades. Clade 8 and clade 6 strains were more frequently associated with severe disease and HUS. In this study, we aimed to identify differentially expressed proteins in two strains of E. coli O157:H7 (clade 8 and clade 6, obtained from cattle and compared them with the well characterized reference EDL933 strain (clade 3. Clade 8 and clade 6 strains show enhanced pathogenicity in a mouse model and virulence-related properties. Proteins were extracted and analyzed using the TMT-6plex labeling strategy associated with two dimensional liquid chromatography and mass spectrometry in tandem. We detected 2241 proteins in the cell extract and 1787 proteins in the culture supernatants. Attention was focused on the proteins related to virulence, overexpressed in clade 6 and 8 strains compared to EDL933 strain. The proteins relevant overexpressed in clade 8 strain were the curli protein CsgC, a transcriptional activator (PchE, phage proteins, Stx2, FlgM and FlgD, a dienelactone hydrolase, CheW and CheY, and the SPATE protease EspP. For clade 6 strain, a high overexpression of phage proteins was detected, mostly from Stx2 encoding phage, including Stx2, flagellin and the protease TagA, EDL933_p0016, dienelactone hydrolase, and Haemolysin A, amongst others with unknown function. Some of these proteins were analyzed by RT-qPCR to corroborate the proteomic data. Clade 6 and clade 8 strains showed enhanced transcription of 10 out of 12 genes compared to EDL933. These results may provide new insights in E. coli O157:H7 mechanisms of pathogenesis.

Characterization of biofilms produced by Escherichia coli O157 isolated from cattle hides

Science.gov (United States)

Milojević, L.; Velebit, B.; Baltić, T.; Nikolić, A.; Mitrović, R.; Đorđević, V.

2017-09-01

This study aimed to investigate possibility E. coli O157 from cattle hides to produced biofilms. We had 28 suspect primoisolates and 17 were confirmed to be E. coli O157. Biofilm production test showed that more than 50% of this isolates did not produce biofilm. From the other half of the isolates, 5 of them were weakly adherent, 3 were moderately adherent. Since E. coli O157 are one of the main foodborne hazards in meat processing industry and the discovery that some of them can produce moderately adherent biofilms, request necessity of strict implementation of HACCP procedures to prevent further expansion this pathogen.

Variation in Resistance of Natural Isolates of Escherichia coli O157 to High Hydrostatic Pressure, Mild Heat, and Other Stresses

Science.gov (United States)

Benito, Amparo; Ventoura, Georgia; Casadei, Maria; Robinson, Tobin; Mackey, Bernard

1999-01-01

Strains of Escherichia coli O157 isolated from patients with clinical cases of food-borne illness and other sources exhibited wide differences in resistance to high hydrostatic pressure. The most pressure-resistant strains were also more resistant to mild heat than other strains. Strain C9490, a representative pressure-resistant strain, was also more resistant to acid, oxidative, and osmotic stresses than the pressure-sensitive strain NCTC 12079. Most of these differences in resistance were observed only in stationary-phase cells, the only exception being acid resistance, where differences were also apparent in the exponential phase. Membrane damage in pressure-treated cells was revealed by increased uptake of the fluorescent dyes ethidium bromide and propidium iodide. When strains were exposed to the same pressure for different lengths of time, the pressure-sensitive strains took up stain sooner than the more resistant strain, which suggested that the differences in resistance may be related to susceptibility to membrane damage. Our results emphasize the importance of including stress-resistant strains of E. coli O157 when the efficacy of a novel or mild food preservation treatment is tested. PMID:10103251

Enterohemorrhagic Escherichia coli O157 in milk and dairy products from Libya: Isolation and molecular identification by partial sequencing of 16S rDNA

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Aboubaker M. Garbaj

2016-11-01

Full Text Available Aim: The aim of this work was to isolate and molecularly identify enterohemorrhagic Escherichia coli (EHEC O157 in milk and dairy products in Libya, in addition; to clear the accuracy of cultural and biochemical identification as compared with molecular identification by partial sequencing of 16S rDNA for the existing isolates. Materials and Methods: A total of 108 samples of raw milk (cow, she-camel, and goat and locally made dairy products (fermented cow’s milk, Maasora, Ricotta and ice cream were collected from some regions (Janzour, Tripoli, Kremiya, Tajoura and Tobruk in Libya. Samples were subjected to microbiological analysis for isolation of E. coli that was detected by conventional cultural and molecular method using polymerase chain reaction and partial sequencing of 16S rDNA. Results: Out of 108 samples, only 27 isolates were found to be EHEC O157 based on their cultural characteristics (Tellurite-Cefixime-Sorbitol MacConkey that include 3 isolates from cow’s milk (11%, 3 isolates from she-camel’s milk (11%, two isolates from goat’s milk (7.4% and 7 isolates from fermented raw milk samples (26%, isolates from fresh locally made soft cheeses (Maasora and Ricotta were 9 (33% and 3 (11%, respectively, while none of the ice cream samples revealed any growth. However, out of these 27 isolates, only 11 were confirmed to be E. coli by partial sequencing of 16S rDNA and E. coli O157 Latex agglutination test. Phylogenetic analysis revealed that majority of local E. coli isolates were related to E. coli O157:H7 FRIK944 strain. Conclusion: These results can be used for further studies on EHEC O157 as an emerging foodborne pathogen and its role in human infection in Libya.

Escherichia coli O157:H7: Animal Reservoir and Sources of Human Infection

Science.gov (United States)

Ferens, Witold A.

2011-01-01

Abstract This review surveys the literature on carriage and transmission of enterohemorrhagic Escherichia coli (EHEC) O157:H7 in the context of virulence factors and sampling/culture technique. EHEC of the O157:H7 serotype are worldwide zoonotic pathogens responsible for the majority of severe cases of human EHEC disease. EHEC O157:H7 strains are carried primarily by healthy cattle and other ruminants, but most of the bovine strains are not transmitted to people, and do not exhibit virulence factors associated with human disease. Prevalence of EHEC O157:H7 is probably underestimated. Carriage of EHEC O157:H7 by individual animals is typically short-lived, but pen and farm prevalence of specific isolates may extend for months or years and some carriers, designated as supershedders, may harbor high intestinal numbers of the pathogen for extended periods. The prevalence of EHEC O157:H7 in cattle peaks in the summer and is higher in postweaned calves and heifers than in younger and older animals. Virulent strains of EHEC O157:H7 are rarely harbored by pigs or chickens, but are found in turkeys. The bacteria rarely occur in wildlife with the exception of deer and are only sporadically carried by domestic animals and synanthropic rodents and birds. EHEC O157:H7 occur in amphibian, fish, and invertebrate carriers, and can colonize plant surfaces and tissues via attachment mechanisms different from those mediating intestinal attachment. Strains of EHEC O157:H7 exhibit high genetic variability but typically a small number of genetic types predominate in groups of cattle and a farm environment. Transmission to people occurs primarily via ingestion of inadequately processed contaminated food or water and less frequently through contact with manure, animals, or infected people. PMID:21117940

[Construction of enterohemorrhagic Escherichia coli O157:H7 strains with espF gene deletion and complementation].

Science.gov (United States)

Hua, Ying; Sun, Qi; Wang, Xiangyu; DU, Yanli; Shao, Na; Zhang, Qiwei; Zhao, Wei; Wan, Chengsong

2015-11-01

To construct enterohemorrhagic Escherichia coli (EHEC) O157:H7 strains with delection espF gene and its nucleotide fragment and with espF gene complementation. A pair of homologous arm primers was designed to amplify the gene fragment of kanamycin resistance, which was transformed into EHEC O157:H7 EDL933w strain via the PKD46 plasmid by electroporation. The replacement of the espF gene by kanamycin resistance gene through the PKD46-mediated red recombination system was confirmed by PCR and sequencing. The entire coding region of espF along with its nucleotide fragment was amplified by PCR and cloned into pBAD33 plasmid, which was transformed into a mutant strain to construct the strain with espF complementation. RT-PCR was used to verify the transcription of espF and its nucleotide fragment in the complemented mutant strain. We established EHEC O157:H7 EDL933w strains with espF gene deletion and with espF gene complementation. Both espF and its nucleotide fragment were transcribed in the complemented mutant strain. The two strains provide a basis for further study of the regulatory mechanism of espF.

Antagonistic Activity of Probiotic Bacteria Isolated from Traditional Dairy Products against E. coli O157:H7

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Maryam Rahimpour Hesari

2017-10-01

Full Text Available Background:    Probiotics are living microorganisms that have useful effects on health of digestive system when acquired in a defined dosage. E. coli O157:H7 is known as one of the most important agents of diarrhea in developing countries. Therefore, attention to the treatment of such gastrointestinal disease is essential. The aim of this study was to determine antagonistic activity of food born lactobacilli against E. coli O157:H7.Methods:     Lactobacilli were isolated from traditional dairy products (yogurt and buttermilk samples. Then, they were characterized using biochemical and molecular tests. Bifidobacterium bifidum PTCC 1644 was obtained from the microbial collection of Iranian Research Organization for Science and Technology in Lyophilized form. Similarly, E. coli O157:H7 PTCC12900 was obtained from faculty of veterinary medicine university of Tehran. The antagonistic activity of probiotics supernatants against E. coli O157:H7 was investigated using the disk diffusion agar, well diffusion agar and pour plate methods.Results:   The isolates were characterized as Lactobacillus plantarum and Lactobacillus fermentum. All isolates showed antagonistic activities against E. coli O157:H7 in all of the three methods, where the activity of L. plantarum and B. bidifum PTCC 1644 was greater than that of L. fermentum. Conclusion:   Metabolites produced by the probiotic bacteria are able to inhibit the growth of E. coli O157:H7. This can be an important solution for the prevention and treatment of E. coli O157:H7 infection and ultimately improve human health.

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.

Isolation, Identification, and Evaluation of Novel Probiotic Strains Isolated from Feces of Breast-Fed Infants.

Science.gov (United States)

Panya, Marutpong; Lulitanond, Viraphong; Rattanachaikunsopon, Pongsak; Srivoramas, Thanyakarn; Chaiwong, Tarinee

2016-01-01

To isolate, identify, and evaluate the probiotic properties of lactic acid bacteria (LAB) isolated from the feces of breast-fed infants. The probiotic tests included investigation of hemolysis activity, survival in simulated gastrointestinal tract conditions (acid and bile salt tolerance), susceptibility to antibiotics, and ability to inhibit selected bacterial pathogens (Escherichia coli O157:H7, Vibrio cholerae and Salmonella enterica subsp enterica serovar Typhimurium). The bacterial species identification was performed by both carbohydrate utilization and partial 16S ribosomal RNA sequencing. Five of fifty LAB isolates (UBU-03, UBU-06, UBU-09, UBU-34, and UBU-37) showed good probiotic properties. These five isolates showed non-hemolysis type (gamma-hemolysis), susceptibility to all antibiotics tested except for vancomycin, ability to survive in the simulated gastrointestinal conditions of both acid and bile salt solution, and ability to inhibit growth of E. coli O157: H7 and V. cholerae. Bacterial species identification revealed that all five isolates were firmly identified as Lactobacillus rhamnosus species. The L. rhamnosus strains that were isolated and characterized in this study could be considered as probiotic strains, and then used for further probiotic characterization in human cell cultures or animal models.

Detection and characterization of Escherichia coli O157:H7 from feral pigeon in Qom province, Iran

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Hossein Esameili

2015-02-01

Full Text Available Objective: To detect and characterize Escherichia coli (E. coli O157:H7 in feral pigeons in Qom province, Iran. Methods: In this survey, 290 cloacal samples were obtained from trapped feral pigeons in Qom province. Microbiological, biochemical and serological examinations were done to detect the E. coli O157:H7. Isolates were subjected to multiplex polymerase chain reaction for the detection of stx1, stx2, eaeA and hlyA genes. Results: Four samples (1.38% were positive for E. coli O157:H7 by using O157 and H7 antisera and only one E. coli O157:H7 strain isolated showed the presence of stx1, stx2, eaeA and hlyA genes. Conclusions: The results of present survey revealed that feral pigeons in Qom province had the potential to be a reservoir of E. coli O157:H7. The low prevalence of E. coli O157:H7 can be attributed to sampling each pigeon just once and fecal culture limits, and true prevalence of the E. coli O157:H7 might be higher than our findings.

[Survival of VTEC O157 and non-O157 in water troughs and bovine feces].

Science.gov (United States)

Polifroni, Rosana; Etcheverría, Analía I; Arroyo, Guillermo H; Padola, Nora L

2014-01-01

Verotoxin-producing Escherichia coli (VTEC) is the etiologic agent of hemolytic-uremic syndrome (HUS), which typically affects children ranging in age from six months to five years old. Transmission is produced by consumption of contaminated food, by direct contact with animals or the environment and from person to person. In previous studies we determined that the environment of a dairy farm is a non-animal reservoir; thus, we proposed to study the survival of 4 VTEC isolates (O20:H19; O91:H21; O157:H7 and O178:H19) in sterile water troughs and bovine feces by viable bacteria count and detection of virulence genes by PCR. It was demonstrated that the survival of different VTEC isolates (O157 and non-O157) varied in terms of their own characteristics as well as of the environmental conditions where they were found. The main differences between isolates were their survival time and the maximal counts reached. The competitive and adaptive characteristics of some isolates increase the infection risk for people that are visiting or working on a farm, as well as the risk for reinfection of the animals and food contamination. Copyright © 2014 Asociación Colombiana de Psiquiatría. Publicado por Elsevier España. All rights reserved.

Comparative sequence analysis revealed altered chromosomal organization and a novel insertion sequence encoding DNA modification and potentially stress-related functions in an Escherichia coli O157:H7 foodborne isolate

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We recently described the complete genome of enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain NADC 6564, an isolate of strain 86-24 linked to the 1986 disease outbreak. In the current study, we compared the chromosomal sequence of NADC 6564 to the well-characterized chromosomal sequences of ...

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

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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.

Characteristics of Shigatoxin-Producing Escherichia coli Strains Isolated during 2010–2014 from Human Infections in Switzerland

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Lisa Fierz

2017-08-01

Full Text Available Objectives: The aim of this study was to characterize a collection of 95 Shigatoxin-producing E.coli (STEC isolated from human patients in Switzerland during 2010–2014.Methods: We performed O and H serotyping and molecular subtyping.Results: The five most common serogroups were O157, O145, O26, O103, and O146. Of the 95 strains, 35 (36.8% carried stx1 genes only, 43 strains (45.2% carried stx2 and 17 (17.9% harbored combinations of stx1 and stx2 genes. Stx1a (42 strains and stx2a (32 strains were the most frequently detected stx subtypes. Genes for intimin (eae, hemolysin (hly, iron-regulated adhesion (iha, and the subtilase cytotoxin subtypes subAB1, subAB2-1, subAB2-2, or subAB2-3 were detected in 70.5, 83.2, 74.7, and 20% of the strains, respectively. Multilocus sequence typing assigned the majority (58.9% of the isolates to five different clonal complexes (CC, 11, 32, 29, 20, and 165, respectively. CC11 included all O157:[H7] and O55:[H7] isolates. CC32 comprised O145:[H28] isolates, and O145:[H25] belonged to sequence type (ST 342. CC29 contained isolates of the O26:[H11], O111:[H8] and O118:[Hnt] serogroups, and CC20 encompassed isolates of O51:H49/[Hnt] and O103:[H2]. CC165 included isolates typed O80:[H2]-ST301, all harboring stx2d, eae-ξ, hly, and 66.7% additionally harboring iha. All O80:[H2]-ST301 strains harbored at least 7 genes carried by pS88, a plasmid associated with extraintestinal virulence. Compared to data from Switzerland from the years 2000–2009, an increase of the proportion of non-O157 STEC infections was observed as well as an increase of infections due to STEC O146. By contrast, the prevalence of the highly virulent German clone STEC O26:[H11]-ST29 decreased from 11.3% during 2000–2009 to 1.1% for the time span 2010–2014. The detection of O80:[H2]-ST301 harboring stx2d, eae-ξ, hly, iha, and pS88 related genes suggests an ongoing emergence in Switzerland of an unusual, highly pathogenic STEC serotype

Evaluation of immunomagnetic separation and PCR for the detection of Escherichia coli O157 in animal feces and meats

NARCIS (Netherlands)

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

2006-01-01

Series of animal feces and meat samples artificially contaminated with strains of Escherichia coli O157 isolated from different sources were tested by both an immunomagnetic separation (IMS)-based method and a PCR method using primers specific for a portion of the rfbE gene of E. coli O157. IMS is

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.

Validation of the detection of STEC (O26, O45, O103, O111, O121, O145 and O157 in raw burgers using real-time PCR (BAX® System Q7, DuPont in “WET POOLS”

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Paula Mussio

2014-12-01

Full Text Available Shiga toxin-producing Escherichia coli (STEC O157:H7 and no-O157:H7 have been identified as emerging foodborne pathogens responsible for an increasing number of outbreaks worldwide. Many foods have been associated to these outbreaks, mainly undercooked beef burgers. Due to the increasing production and consumption of burgers in our country, it is important to have a rapid technique to identify and isolate the most important STEC strains in foods matrixes.The objective of these investigation was to assess the sensitivity, specificity and limit detection for the screening of the most prevalent STEC in frozen raw beef burgers, using the "STEC Screening stx/eae " kit to detect the stx/eae genes and the "Panel 1 STEC E. coli O26, O111, O121" and "Panel 2 STEC E. coli O45, O103, O145" and "E. coli O157: H7 MP” (DuPont kits for the detection of the different serogroups. The use of composed samples (wet pools, the recovery of each strain from the positives samples on selective media after specified inmunoconcentration and the detection of stx/eae virulence genes in isolates by PCR were also evaluated. We validated a technique that allows the detection of the mentioned STEC strains with limits of detection between 1-5 CFU in 65 grams of raw frozen hamburgers, using wet pools. 

Prevalence and Genomic Characterization of Escherichia coli O157:H7 in Cow-Calf Herds throughout California.

Science.gov (United States)

Worley, Jay N; Flores, Kristopher A; Yang, Xun; Chase, Jennifer A; Cao, Guojie; Tang, Shuai; Meng, Jianghong; Atwill, Edward R

2017-08-15

Escherichia coli serotype O157:H7 is a zoonotic food- and waterborne bacterial pathogen that causes a high hospitalization rate and can cause life-threatening complications. Increasingly, E. coli O157:H7 infections appear to originate from fresh produce. Ruminants, such as cattle, are a prominent reservoir of E. coli O157:H7 in the United States. California is one of the most agriculturally productive regions in the world for fresh produce, beef, and milk. The close proximity of fresh produce and cattle presents food safety challenges on a uniquely large scale. We performed a survey of E. coli O157:H7 on 20 farms in California to observe the regional diversity and prevalence of E. coli O157:H7. Isolates were obtained from enrichment cultures of cow feces. Some farms were sampled on two dates. Genomes from isolates were sequenced to determine their relatedness and pathogenic potential. E. coli O157:H7 was isolated from approximately half of the farms. The point prevalence of E. coli O157:H7 on farms was highly variable, ranging from zero to nearly 90%. Within farms, generally one or a few lineages were found, even when the rate of isolation was high. On farms with high isolation rates, a single clonal lineage accounted for most of the isolates. Farms that were visited months after the first visit might have had the same lineages of E. coli O157:H7. Strains of E. coli O157:H7 may be persistent for months on farms. IMPORTANCE This survey of 20 cow-calf operations from different regions of California provides an in depth look at resident Escherichia coli O157:H7 populations at the molecular level. E. coli O157:H7 is found to have a highly variable prevalence, and with whole-genome sequencing, high prevalences in herds were found to be due to a single lineage shed from multiple cows. Few repeat lineages were found between farms in this area; therefore, we predict that E. coli O157:H7 has significant diversity in this area beyond what is detected in this survey. All

Prevalence and antibiogram profiles of Escherichia coli O157:H7 isolates recovered from three selected dairy farms in the Eastern Cape Province, South Africa

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Luyanda Msolo

2016-12-01

Full Text Available Objective: To investigate the occurrence and antibiotics susceptibility of Escherichia coli (E. coli O157:H7 isolates from raw milk, cattle udder, milking machines and worker’s hand swabs from three selected commercial dairy farms in the Amathole District Municipality, Eastern Cape Province, South Africa. Methods: Raw milk samples were collected from bulk storage tanks and swab samples were collected from milking machines, cattle udders and worker’s hands fortnightly over a sixmonth sampling regime between June and November 2014. A standard culture-based method was used for the enumeration and isolation of E. coli O157:H7, presumptive identification using sorbitol MacConkey agar (supplemented with cefixime (50 µg/L and potassium tellurite (25 mg/L. A serological confirmation of the presumptive E. coli O157:H7 isolates was conducted using the O157 latex agglutination test kit. Results: A total of 252 E. coli O157:H7 isolates were further subjected to PCR amplification of rfbEO157 and flCH7 genes of which 27(11% of the isolates were confirmed positive E. coli O157:H7. The percentage antibiotic resistance of the 27 E. coli O157:H7 isolates from the dairy farms revealed penicillin [23 (85%], tetracycline [22 (81%], erythromycin [19 (70%], streptomycin [14 (52%] and chloramphenicol [12 (45%]. The highest resistances were penicillin [23 (85%] and tetracycline [22 (81%]. Conclusions: These findings revealed that the dairy farms are potential reservoirs of E. coli O157:H7 serotype, and harbor antibiotic-resistant determinants, a concern to public and environmental health.

Preparation of immunomagnetic iron-dextran nanoparticles and application in rapid isolation of E.coli O157:H7 from foods

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Duan, Hui-Li; Shen, Zhi-Qiang; Wang, Xin-Wei; Chao, Fu-Huan; Li, Jun-Wen

2005-01-01

AIM: To prepare a kind of magnetic iron-dextran nanoparticles that was coated with anti-E.coli O157:H7 IgG, analyze its application conditions, and try to use it to isolate E.coli O157:H7 from foods. METHODS: Magnetic iron-dextran nanoparticles were prepared by the reaction of a mixture of ferric and ferrous ions with dextran polymers under alkaline conditions. The particles were coated with antiserum against E.coli O157:H7 by the periodate oxidation-borohydride reduction procedure. The oxidation time, amount of antibody coating the particles, amount of nanoparticles, incubation time and isolation time were varied to determine their effects on recovery of the organisms. Finally, the optimum conditions for isolating E.coli O157:H7 from food samples were established. RESULTS: E.coli O157:H7 can be isolated from samples within 15 min with the sensitivity of 101 CFU/mL or even less. In the presence of 108 CFU/mL of other organisms, the sensitivity is 101-102 CFU/mL. Nonspecific binding of other bacteria to the particles was not observed. Two and a half hours of enrichment is enough for the particles to detect the target from the food samples inoculated with 1 CFU/g. CONCLUSION: Isolation of target bacteria by immuno-magnetic nanoparticles is an efficient method with high sensitivity and specificity. The technique is so simple that it can be operated in lab and field even by untrained personnel. PMID:15968716

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.

Elimination of Pathogen Escherichia coli O157:H7 in Ground Beef by a Newly Isolated Strain of Lactobacillus acidophilus during Storage at 5°C

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Alireza Goodarzi

2016-06-01

Full Text Available Background and Objective: Constant use of limited number of lactic acid bacteria species in biopreservation can cause genetic degradation and or rising resistance against food pathogens or antimicrobial substances they produce. For this objective, a newly isolated strain of Lactobacillus acidophilus possessing high antimicrobial activity was evaluated as a candidate for use in biopreservation.Materials and Methods: Antibacterial activity was evaluated by agar disk diffusion method. Hydrogen peroxide amount was measured by Merckoquant Peroxide test strips. Microbiological analysis of the ground beef infected by Escherichia coli O157:H7 and treated by Lactobacillus acidophilus GH 201was done by plating of serial dilution in physiological saline on Tryptose agar.Results and Conclusion: The cells (109 CFU ml-1 of Lactobacillus acidophilus produced significant amount of antibacterial substances mainly hydrogen peroxide (28 and 30 μg ml-1 in sodium phosphate buffer (0.2 M, pH 6.5 and LAPTg at 5°C during submerged cultivation with no growth, respectively. Submerged co-cultivation of Escherichia coli O157:H7 with lactobacilli in LAPTg broth at 5°C reduced the total number of the pathogen more than 2 log for 5 days. In case of solid state cultivation on agar-based medium, the maximum inhibitory zones on Escherichia coli O157:H7 lawn around the disks soaked by different amounts of washed Lactobacillus acidophilus cells appear for one-day cold exposition. The size of inhibition zone depends on the concentration of lactic acid bacteria cells. The cell suspension intended for treatment must contain 108-9CFU ml-1 of lactic acid bacteria. Lactobacillus acidophilus reduced the initial amount (2×105 CFU ml-1 of Escherichia coli O157:H7 in ground beef up to 2 log for 5 days of solid-state co-cultivation. The application of Lactobacillus acidophilus bacteria expanded the shelf-life of ground beef due to inhibition of

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

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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

E.coli O157:H7

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Treor Francis Fernandez

2008-06-01

Full Text Available The serious nature of the symptoms of haemorrhagic colitis and HUS and the apparent low infectious dose (<100 cells of E.coli O157:H7 places this food borne pathogen a most serious of known food borne pathogens. Persuasive evidence suggests that healthy cattle are a reservoir of O157 and they can enter the food chain to provide a source of exposure for humans. A possible route of transmission of O157 VTEC may involve infections initially in calves that shed their organism into faecal slurry that may be used on grazing grass. This provides potential for infection of other animals from which the organism may contaminate milk or carcasses at slaughter. Possible sources of VTEC in healthy animals other E.coli O157:H7 than cattle and a wider range of foodstuffs require further investigation. Many features of E.coli O157:H7 strains remain poorly understood. It includes: (i Role of virulent genes in the animal, (ii Mechanism of evolution of the organism, (iii The progress of individual cases of E.coli O157:H7 infection, and (iv The difference in incidence of infection in different geographical areas. [Veterinary World 2008; 1(3.000: 83-87

Biological characteristics and probiotic effect of Leuconostoc lactis strain isolated from the intestine of black porgy fish

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Wei Zhang

2013-09-01

Full Text Available A strain of lactic acid bacteria, Leuconostoc lactis, was isolated from the intestinal tract of black porgy, Sparus macrocephalus, and identified by conventional biochemical characteristics and 16S rDNA gene sequence analysis. The isolated strain had the ability of bile tolerance and resistance to low pH, and survived well in the trypsinase and pepsin solution. But the highly concentrated dose of trypsinase and pepsin affect the viability of the isolated strain. The isolate was resistant to several antibiotics, including Cephalothin, Ceftriaxone, Imipenem and Tobramycin. The isolate could autoaggregate itself and coaggregate with other bacteria in vitro. The autoaggregation percentage increased to 23.29% after 20 h of incubation. The percentage of coaggregation were respectively 31.21%, 29.44%, 10.74%, 16.49%, 24.36%, 24.41% and 20.99% for Vibrio parahaemolyticus, Listeria monocytogenes, Escherichia coli O157, Salmonella typhimurium, Shigella, Staphylococcus aureus and Proteusbacillus vulgaris after 20 h incubation of a mixed suspension. The supernatant of the strain inhibited the growth of several pathogens, such as V.parahaemolyticus, Vibrio harveyi, Vibrio alginolyticus, Staphylococcus aureus, Escherichia coli O157, Salmonella typhimurium, Bacillus subtilis, Proteusbacillus vulgaris and Shigella. These results indicated that the isolate, Leuconostoc lactis, might be an attractive candidate for perspectival strain for probiotics in marine aquaculture.

Foodborne transmission of sorbitol-fermenting Escherichia coli O157:[H7] via ground beef: an outbreak in northern France, 2011.

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King, L A; Loukiadis, E; Mariani-Kurkdjian, P; Haeghebaert, S; Weill, F-X; Baliere, C; Ganet, S; Gouali, M; Vaillant, V; Pihier, N; Callon, H; Novo, R; Gaillot, O; Thevenot-Sergentet, D; Bingen, E; Chaud, P; de Valk, H

2014-12-01

Sorbitol-fermenting Escherichia coli O157:[H7] is a particularly virulent clone of E. coli O157:H7 associated with a higher incidence of haemolytic uraemic syndrome and a higher case fatality rate. Many fundamental aspects of its epidemiology remain to be elucidated, including its reservoir and transmission routes and vehicles. We describe an outbreak of sorbitol-fermenting E. coli O157:[H7] that occurred in France in 2011. Eighteen cases of paediatric haemolytic uraemic syndrome with symptom onset between 6 June and 15 July 2011 were identified among children aged 6 months to 10 years residing in northern France. A strain of sorbitol-fermenting E. coli O157:[H7] stx2a eae was isolated from ten cases. Epidemiological, microbiological and trace-back investigations identified multiply-contaminated frozen ground beef products bought in a supermarket chain as the outbreak vehicle. Strains with three distinct pulsotypes that were isolated from patients, ground beef preparations recovered from patients' freezers and from stored production samples taken at the production plant were indistinguishable upon molecular comparison. This investigation documents microbiologically confirmed foodborne transmission of sorbitol-fermenting of E. coli O157 via beef and could additionally provide evidence of a reservoir in cattle for this pathogen. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

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.

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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.

Severe Outbreak of Sorbitol-Fermenting Escherichia coli O157 via Unpasteurized Milk and Farm Visits, Finland 2012.

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Jaakkonen, A; Salmenlinna, S; Rimhanen-Finne, R; Lundström, H; Heinikainen, S; Hakkinen, M; Hallanvuo, S

2017-09-01

Shiga toxin-producing, sorbitol-fermenting Escherichia coli O157 (SF O157) has emerged as a cause of severe human illness. Despite frequent human findings, its transmission routes and reservoirs remain largely unknown. Foodborne transmission and reservoir in cattle have been suspected, but with limited supporting evidence. This study describes the outbreak of SF O157 that occurred in Finland in 2012. The outbreak originated from a recreational farm selling unpasteurized milk, as revealed by epidemiologic and microbiological investigations, and involved six hospitalized children and two asymptomatic adults with culture-confirmed infection. An identical strain of SF O157 was isolated from patients, cattle and the farm environment, and epidemiologic analysis suggested unpasteurized milk as the vehicle of transmission. This study reports the first milkborne outbreak of SF O157, provides supporting evidence of cattle as a reservoir and highlights the health risks related to the consumption of unpasteurized milk. © 2017 The Authors. Zoonoses and Public Health Published by Blackwell Verlag GmbH.

Comparison of clinical and immunological findings in gnotobiotic piglets infected with Escherichia coli O104:H4 outbreak strain and EHEC O157:H7.

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Wöchtl, Bettina; Gunzer, Florian; Gerner, Wilhelm; Gasse, Hagen; Koch, Michaela; Bagó, Zoltán; Ganter, Martin; Weissenböck, Herbert; Dinhopl, Nora; Coldewey, Sina M; von Altrock, Alexandra; Waldmann, Karl-Heinz; Saalmüller, Armin; Zimmermann, Kurt; Steinmann, Jörg; Kehrmann, Jan; Klein-Hitpass, Ludger; Blom, Jochen; Ehricht, Ralf; Engelmann, Ines; Hennig-Pauka, Isabel

2017-01-01

Shiga toxin (Stx) producing Escherichia coli ( E. coli ) (STEC) is the most frequent cause of diarrhoea-positive haemolytic uraemic syndrome (D + HUS) in humans. In 2011, a huge outbreak with an STEC O104:H4 strain in Germany highlighted the limited possibilities for causative treatment of this syndrome. The responsible STEC strain was found to combine Stx production with adherence mechanisms normally found in enteroaggregative E. coli (EAEC). Pathotypes of E. coli evolve and can exhibit different adhesion mechanisms. It has been shown previously that neonatal gnotobiotic piglets are susceptible for infection with STEC, such as STEC O157:H7 as well as for EAEC, which are considered to be the phylogenetic origin of E. coli O104:H4. This study was designed to characterise the host response to infection with the STEC O104:H4 outbreak strain in comparison to an STEC O157:H7 isolate by evaluating clinical parameters (scoring) and markers of organ dysfunction (biochemistry), as well as immunological (flow cytometry, assessment of cytokines/chemokines and acute phase proteins) and histological alterations (light- and electron microscopy) in a gnotobiotic piglet model of haemolytic uraemic syndrome. We observed severe clinical symptoms, such as diarrhoea, dehydration and neurological disorders as well as attaching-and-effacing lesions (A/E) in the colon in STEC O157:H7 infected piglets. In contrast, STEC O104:H4 challenged animals exhibited only mild clinical symptoms including diarrhoea and dehydration and HUS-specific/severe histopathological, haematological and biochemical alterations were only inconsistently presented by individual piglets. A specific adherence phenotype of STEC O104:H4 could not be observed. Flow cytometric analyses of lymphocytes derived from infected animals revealed an increase of natural killer cells (NK cells) during the course of infection revealing a potential role of this subset in the anti-bacterial activity in STEC disease. Unexpectedly, E

Genome sequences of thirty Escherichia coli O157:H7 isolates recovered from a single dairy farm and its associated off-site heifer raising facility

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Cattle are the primary reservoir of Escherichia coli O157:H7, the most frequently isolated serotype of enterohemorrhagic E. coli infections among humans in North America. To evaluate the diversity of E. coli O157:H7 isolates within a single dairy herd the genomes of 30 isolates collected over a 7-ye...

Green fluorescent protein labeling of Listeria, Salmonella, and Escherichia coli O157:H7 for safety-related studies.

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Li Ma

Full Text Available Many food safety-related studies require tracking of introduced foodborne pathogens to monitor their fate in complex environments. The green fluorescent protein (GFP gene (gfp provides an easily detectable phenotype so has been used to label many microorganisms for ecological studies. The objectives of this study were to label major foodborne pathogens and related bacteria, including Listeria monocytogenes, Listeria innocua, Salmonella, and Escherichia coli O157:H7 strains, with GFP and characterize the labeled strains for stability of the GFP plasmid and the plasmid's effect on bacterial growth. GFP plasmids were introduced into these strains by a CaCl(2 procedure, conjugation or electroporation. Stability of the label was determined through sequential propagation of labeled strains in the absence of selective pressure, and rates of plasmid-loss were calculated. Stability of the GFP plasmid varied among the labeled species and strains, with the most stable GFP label observed in E. coli O157:H7. When grown in nonselective media for two consecutive subcultures (ca. 20 generations, the rates of plasmid loss among labeled E. coli O157:H7, Salmonella and Listeria strains ranged from 0%-30%, 15.8%-99.9% and 8.1%-93.4%, respectively. Complete loss (>99.99% of the plasmid occurred in some labeled strains after five consecutive subcultures in the absence of selective pressure, whereas it remained stable in others. The GFP plasmid had an insignificant effect on growth of most labeled strains. E. coli O157:H7, Salmonella and Listeria strains can be effectively labeled with the GFP plasmid which can be stable in some isolates for many generations without adversely affecting growth rates.

Fitness of Enterohemorrhagic Escherichia coli (EHEC)/Enteroaggregative E. coli O104:H4 in Comparison to That of EHEC O157: Survival Studies in Food and In Vitro.

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Böhnlein, Christina; Kabisch, Jan; Meske, Diana; Franz, Charles M A P; Pichner, Rohtraud

2016-11-01

In 2011, one of the world's largest outbreaks of hemolytic-uremic syndrome (HUS) occurred, caused by a rare Escherichia coli serotype, O104:H4, that shared the virulence profiles of Shiga toxin-producing E. coli (STEC)/enterohemorrhagic E. coli (EHEC) and enteroaggregative E. coli (EAEC). The persistence and fitness factors of the highly virulent EHEC/EAEC O104:H4 strain, grown either in food or in vitro, were compared with those of E. coli O157 outbreak-associated strains. The log reduction rates of the different EHEC strains during the maturation of fermented sausages were not significantly different. Both the O157:NM and O104:H4 serotypes could be shown by qualitative enrichment to be present after 60 days of sausage storage. Moreover, the EHEC/EAEC O104:H4 strain appeared to be more viable than E. coli O157:H7 under conditions of decreased pH and in the presence of sodium nitrite. Analysis of specific EHEC strains in experiments with an EHEC inoculation cocktail showed a dominance of EHEC/EAEC O104:H4, which could be isolated from fermented sausages for 60 days. Inhibitory activities of EHEC/EAEC O104:H4 toward several E. coli strains, including serotype O157 strains, could be determined. Our study suggests that EHEC/EAEC O104:H4 is well adapted to the multiple adverse conditions occurring in fermented raw sausages. Therefore, it is strongly recommended that STEC strain cocktails composed of several serotypes, instead of E. coli O157:H7 alone, be used in food risk assessments. The enhanced persistence of EHEC/EAEC O104:H4 as a result of its robustness, as well as the production of bacteriocins, may account for its extraordinary virulence potential. In 2011, a severe outbreak caused by an EHEC/EAEC serovar O104:H4 strain led to many HUS sequelae. In this study, the persistence of the O104:H4 strain was compared with those of other outbreak-relevant STEC strains under conditions of fermented raw sausage production. Both O157:NM and O104:H4 strains could survive

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

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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.

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.

Identification of protozoa in dairy lagoon wastewater that consume Escherichia coli O157:H7 preferentially.

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Subbarao V Ravva

Full Text Available Escherichia coli O157:H7 (EcO157, an agent of life threatening hemolytic-uremic syndrome, resides in ruminants and is released in feces at numbers as high as 10 million cells/gram. EcO157 could survive in manure for as long as 21 months, but we observed a 90% decrease in cells of an outbreak strain of EcO157 within half a day in wastewater from dairy lagoons. Although chemical, environmental and biological factors may be responsible for this decrease, we observed an 11-fold increase in native protozoa when wastewater was re-inoculated with 2×10(7 cells of EcO157/mL. These protozoa engulfed the green fluorescent protein labeled EcO157 within 2 hours after inoculation, but expelled vacuoles filled with live EcO157 cells within 3 days into surrounding wastewater, whereas other protozoa retained the EcO157-filled vacuoles for 7 days. EcO157 was not detected by confocal microscopy either inside or outside protozoa after 7 days. Mixed cultures of protozoa enriched from wastewater consumed EcO157 preferentially as compared to native aerobic bacteria, but failed to eliminate them when EcO157 cells declined to 10(4/mL. We isolated three protozoa from mixed cultures and typed them by 18S sequencing as Vorticella microstoma, Platyophyra sp. and Colpoda aspera. While all three protozoa internalized EcO157, only Platyophyra and Colpoda acted as predators. Similar to mixed cultures, these protozoa failed to eliminate EcO157 from PBS containing no other supplemental nutrients or prey. However, spiking PBS with cereal grass medium as nutrients induced predation of EcO157 by Platyophyra sp. after 3 days or enhanced predation by Colpoda after 5 days. Therefore, attempts to enrich protozoa to decrease EcO157 from dairy lagoons, may correspond to an increase in protozoa similar to Vorticella and possibly facilitate transport of bacterial pathogens to food crops grown in proximity.

Co-ordinate single-cell expression of LEE4- and LEE5-encoded proteins of Escherichia coli O157:H7.

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Roe, Andrew J; Naylor, Stuart W; Spears, Kevin J; Yull, Helen M; Dransfield, Tracy A; Oxford, Matthew; McKendrick, Iain J; Porter, Megan; Woodward, Martin J; Smith, David G E; Gally, David L

2004-10-01

Escherichia coli O157:H7 is a zoonotic pathogen that can express a type III secretion system (TTSS) considered important for colonization and persistence in ruminants. E. coli O157:H7 strains have been shown to vary markedly in levels of protein secreted using the TTSS and this study has confirmed that a high secretion phenotype is more prevalent among isolates associated with human disease than isolates shed by healthy cattle. The variation in secretion levels is a consequence of heterogeneous expression, being dependent on the proportion of bacteria in a population that are actively engaged in protein secretion. This was demonstrated by indirect immunofluorescence and eGFP fusions that examined the expression of locus of enterocyte effacement (LEE)-encoded factors in individual bacteria. In liquid media, the expression of EspA, tir::egfp, intimin, but not map::egfp were co-ordinated in a subpopulation of bacteria. In contrast to E. coli O157:H7, expression of tir::egfp in EPEC E2348/69 was equivalent in all bacteria although the same fusion exhibited variable expression when transformed into an E. coli O157:H7 background. An E. coli O157:H7 strain deleted for the LEE demonstrated weak but variable expression of tir::egfp indicating that the elements controlling the heterogeneous expression lie outside the LEE. The research also demonstrated the rapid induction of tir::egfp and map::egfp on contact with bovine epithelial cells. This control in E. coli O157:H7 may be required to limit exposure of key surface antigens, EspA, Tir and intimin during colonization of cattle but allow their rapid production on contact with bovine gastrointestinal epithelium at the terminal rectum.

Inactivation of Escherichia coli O157:H7 on stainless steel upon exposure to Paenibacillus polymyxa biofilms.

Science.gov (United States)

Kim, Seonhwa; Bang, Jihyun; Kim, Hoikyung; Beuchat, Larry R; Ryu, Jee-Hoon

2013-11-01

We investigated the potential use of biofilm formed by a competitive-exclusion (CE) microorganism to inactivate Escherichia coli O157:H7 on a stainless steel surface. Five microorganisms showing inhibitory activities against E. coli O157:H7 were isolated from vegetable seeds and sprouts. The microorganism with the greatest antimicrobial activity was identified as Paenibacillus polymyxa (strain T5). In tryptic soy broth (TSB), strain T5 reached a higher population at 25 °C than at 12 or 37 °C without losing inhibitory activity against E. coli O157:H7. When P. polymyxa (6 log CFU/mL) was co-cultured with E. coli O157:H7 (2, 3, 4, or 5 log CFU/mL) in TSB at 25 °C, the number of E. coli O157:H7 decreased significantly within 24h. P. polymyxa formed a biofilm on stainless steel coupons (SSCs) in TSB at 25 °C within 24h, and cells in biofilms, compared to attached cells without biofilm formation, showed significantly increased resistance to a dry environment (43% relative humidity [RH]). With the exception of an inoculum of 4 log CFU/coupon at 100% RH, upon exposure to biofilm formed by P. polymyxa on SSCs, populations of E. coli O157:H7 (2, 4, or 6 log CFU/coupon) were significantly reduced within 48 h. Most notably, when E. coli O157:H7 at 2 log CFU/coupon was applied to SSCs on which P. polymyxa biofilm had formed, it was inactivated within 1h, regardless of RH. These results will be useful when developing strategies using biofilms produced by competitive exclusion microorganisms to inactivate foodborne pathogens in food processing environments. © 2013.

Temperature-Dependent Fermentation of d-Sorbitol in Escherichia coli O157:H7

OpenAIRE

Bouvet, O. M. M.; Pernoud, S.; Grimont, P. A. D.

1999-01-01

The influence of growth temperature on the ability to ferment d-sorbitol was investigated in Escherichia coli O157:H7. It was found that O157:H7 strains have a temperature-sensitive sorbitol phenotype. d-Sorbitol transport and sorbitol-6-phosphate dehydrogenase activities were expressed in sorbitol-fermenting cells grown at 30°C but only at a low level at 40°C. Sorbitol-positive variants able to transport d-sorbitol were easily selected at 30°C from culture of Sor− E. coli O157:H7 strains.

Lessons learned from a textbook outbreak: EHEC-O157:H7 infections associated with the consumption of raw meat products, June 2012, Limburg, Belgium.

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Braeye, Toon; Denayer, Sarah; De Rauw, Klara; Forier, Anmarie; Verluyten, Jurgen; Fourie, Ludo; Dierick, Katelijne; Botteldoorn, Nadine; Quoilin, Sophie; Cosse, Pascale; Noyen, Jeannine; Pierard, Denis

2014-01-01

On 5 June 2012 several enterohemorrhagic Escherichia coli, EHEC, O157:H7 infections were reported to the public health authorities of Limburg. We performed a case-control study, a trace back/forward investigation and compared strains isolated from human cases and food samples. A case was defined as anyone with a laboratory-confirmed E. coli O157:H7-infection in North-East Limburg from May 30 2012 till July 15 2012. Family members with bloody diarrhea were also included as cases. E. coli O157 was isolated by culture and the presence of the virulence genes was verified using (q)PCR. Isolates were genotyped and compared by Pulsed Field Gel Electrophoresis (PFGE) and insertion sequence 629-printing (IS629-printing). The outbreak involved 24 cases, of which 17 were laboratory-confirmed. Five cases developed Hemolytic Uremic Syndrome (HUS) and fifteen were hospitalized. Cases reported a significantly higher consumption of "steak tartare", a raw meat product (OR 48.12; 95% CI; 5.62- 416.01). Cases were also more likely to buy meat-products at certain butcheries (OR 11.67; 95% CI; 1.41 - 96.49). PFGE and IS629-printing demonstrated that the vtx1a vtx2a eae ehxA positive EHEC O157:H7 strains isolated from three meat products and all seventeen human stool samples were identical. In a slaughterhouse, identified by the trace-back investigation, a carcass infected with a different EHEC strain was found and confiscated. We present a well described and effectively investigated foodborne outbreak associated with meat products. Our main recommendations are the facilitation and acceleration of the outbreak detection and the development of a communication plan to reaches all persons at risk. Foodborne diseases, Shiga-toxigenic Escherichia coli, Enterohemorrhagic Escherichia coli, Meat products, Case control studies, Electrophoresis, Gel, Pulsed-Field.

Investigation of carbon storage regulation network (csr genes) and phenotypic differences between acid sensitive and resistant Escherichia coli O157:H7 strains

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Background: Escherichia coli O157:H7 and related serotype strains have previously been shown to vary in acid resistance, however, little is known about strain specific mechanisms of acid resistance. We examined sensitive and resistant E. coli strains to determine the effects of growth in minimal and...

Screening of the novel colicinogenic gram-negative rods against pathogenic Escherichia coli O157:H7

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H Mushtaq

2015-01-01

Full Text Available Purpose: Escherichia coli (E. coli O157:H7 is gram-negative enteric pathogen producing different types of Shiga toxin. This bacterium is the most corporate cause of haemorrhagic colitis in human. Administration of antibiotics (particularly sulfa drugs against this pathogen is a debatable topic as this may increase the risk of uremic syndrome; especially in children and aged people. Around the world, microbiologists are in search of alternative therapeutic methods specially probiotics against this pathogen. In the present study, we have focused on the investigation of alternate bio-therapeutics (probiotics for the treatment of patients infected with E. coli O157:H7. This study is based on the identification of colicin-producing gram-negative bacteria (particularly enterobacteriaceae which can competently exclude E. coli O157:H7 from the gut of the infected individual. Materials and Methods: Hundred samples from human, animal faeces and septic tank water were analysed for nonpathogenic gram-negative rods (GNRs. Results: Out of these samples, 175 isolates of GNRs were checked for their activity against E. coli O157:H7. Only 47 isolates inhibited the growth of E. coli O157:H7, among which majority were identified as E. coli. These E. coli strains were found to be the efficient producers of colicin. Some of the closely related species i. e., Citrobacter sp, Pantoea sp. and Kluyvera sp. also showed considerable colicinogenic activity. Moreover, colicinogenic species were found to be nonhaemolytic, tolerant to acidic environment (pH 3 and sensitive to commonly used antibiotics. Conclusion: Nonhaemolytic, acid tolerant and sensitive to antibiotics suggests the possible use of these circulating endothelial cells (CEC as inexpensive and inoffensive therapeutic agent (probiotics in E. coli O157:H7 infections.

Sources of Escherichia coli O157 and experiences over the past 15 years in Sheffield, UK.

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Chapman, P A

2000-01-01

In the first documented outbreak of HC caused by Escherichia coli O157, which occurred in the North-west USA in 1982, there was a strong association between infection and prior consumption of ground beef from a chain of fast food restaurants. Foods of bovine origin, including beef, milk and dairy products, have since been implicated in many outbreaks of infection world-wide. Investigations during the course of outbreaks, or at random, have shown that cattle are a major reservoir of E. coli O157. E. coli O157 was isolated from cattle at slaughter in Sheffield in 1987, this being the first isolation from cattle in the UK. Following a cluster of cases in May/June 1992, an abattoir study showed the organism to be present in 4% of cattle at slaughter and on up to a third of carcasses from rectal swab-positive animals. E. coli O157 was isolated from a food source (unpasteurized milk), for the first time in the UK, in Sheffield in May 1993. During surveillance in 1995-6, E. coli O157 was isolated from 15.7% of cattle, with a monthly prevalence which varied from 5 to 37%. E. coli O157 was also isolated from 2.2% of sheep. During surveillance in 1996, E. coli O157 was isolated from 5.9% of samples of lamb products and from 1.5% of samples of beef products, despite the prevalence in cattle being much higher than in sheep. Work is in progress to try to explain this higher prevalence in lamb products. During 1997 in Sheffield, the only cases of E. coli O157 for which a confirmed source was established were associated with direct animal contact on farm visits. During on-farm investigations of these cases, E. coli O157 was isolated from faecal samples from adult cattle, calves, three different breeds of sheep, two different breeds of pigs, goats and a pony.

Cloacael Carriage and Multidrug Resistance Escherichia coli O157:H7 from Poultry Farms, Eastern Ethiopia.

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Shecho, Mude; Thomas, Naod; Kemal, Jelalu; Muktar, Yimer

2017-01-01

A cross-sectional study was carried out to determine antimicrobial drug resistance patterns of E. coli O157:H7 isolates and estimate the level of the pathogen. A total of 194 cloacae swab samples were collected randomly in two poultry farms. Standard cultural, biochemical, and serological (latex agglutination) methods were used to isolate E. coli O157:H7. The isolates were subjected to antimicrobial susceptibility testing using disc diffusion method. Out of 194 cloacae samples examined, 13.4% ( n = 26) were found to be positive for E. coli O157:H7. The finding indicated differences in E. coli O157:H7 infection among the different risk factors. Chicken from Adele Poultry Farm showed higher E. coli O157:H7 infection (OR = 3.89) than Haramaya University poultry farm and young birds had more infection (OR = 4.62) than adult birds. Of the total 14 antimicrobials included in the panel of study, the susceptibility results were varied with 96.15% and 0% E. coli O157:H7 isolates expressing resistance to erythromycin, clindamycin, spectinomycin, and ciprofloxacin, respectively. Multidrug resistance to more than two antimicrobial agents was detected in 24 (92.30%) of the isolates. The study showed high presence of antimicrobial resistant isolates of E. coli O157:H7. Further study is required to better understand the ecology and evolution of bacterial resistance to antimicrobial agents.

Cloacael Carriage and Multidrug Resistance Escherichia coli O157:H7 from Poultry Farms, Eastern Ethiopia

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Mude Shecho

2017-01-01

Full Text Available A cross-sectional study was carried out to determine antimicrobial drug resistance patterns of E. coli O157:H7 isolates and estimate the level of the pathogen. A total of 194 cloacae swab samples were collected randomly in two poultry farms. Standard cultural, biochemical, and serological (latex agglutination methods were used to isolate E. coli O157:H7. The isolates were subjected to antimicrobial susceptibility testing using disc diffusion method. Out of 194 cloacae samples examined, 13.4% (n=26 were found to be positive for E. coli O157:H7. The finding indicated differences in E. coli O157:H7 infection among the different risk factors. Chicken from Adele Poultry Farm showed higher E. coli O157:H7 infection (OR = 3.89 than Haramaya University poultry farm and young birds had more infection (OR = 4.62 than adult birds. Of the total 14 antimicrobials included in the panel of study, the susceptibility results were varied with 96.15% and 0% E. coli O157:H7 isolates expressing resistance to erythromycin, clindamycin, spectinomycin, and ciprofloxacin, respectively. Multidrug resistance to more than two antimicrobial agents was detected in 24 (92.30% of the isolates. The study showed high presence of antimicrobial resistant isolates of E. coli O157:H7. Further study is required to better understand the ecology and evolution of bacterial resistance to antimicrobial agents.

Incidence and antimicrobial susceptibility of Escherichia coli O157:H7 isolates recovered from dairy farms in Amathole District Municipality, Eastern Cape, South Africa

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Asive Myataza

2017-11-01

Full Text Available Objective: To assess the incidence of Escherichia coli (E. coli O157:H7 in water and cattle rectal samples from three commercial dairy farms in Amathole District Municipalities in the Eastern Cape Province of South Africa. Methods: Samples were collected bimonthly from cattle rectum and dairy water sources including irrigation water, drinking water troughs and wastewater between June and November 2014. Standard culture-based methods were applied for the microbial analyses, the disc diffusion method was employed for the antibiotic susceptibility test and PCR approach was utilized for identification of the isolates. Results: A total of 252 presumptive E. coli O157:H7 were isolated and subjected to molecular confirmation by PCR. About 18.7% (47/252 of these were confirmed as E. coli O157:H7. The antimicrobial susceptibility profile of these confirmed isolates revealed high-level resistance against penicillin G (81%, tetracycline (43%, oxytetracycline (62%, erythromycin (68%, sulphamethoxazole (57%, chloramphenicol (55%, doxycycline (51% and trimethoprimsulphamethoxazole (45%. Conclusions: This is the first report of multi-drug resistance E. coli O157:H7 in commercial dairy farms in the province and suggests the possibility of same in other provinces of the country, and this is the subject of the intensive investigation in our group.

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.

Survival of Antibiotic Resistant and Antibiotic Sensitive Strains of E. coli O157 and E. coli O26 in Food Matrices.

OpenAIRE

Walsh, Ciara; Duffy, Geraldine; Blair, I. S.; McDowell, D. A.

2006-01-01

Escherichia coli O157:H7 or E. coli O26, which were AS (antibiotic sensitive), AR (laboratory created antibiotic resistant mutants), or naturally MAR (multi-antibiotic resistant), were inoculated into laboratory media, yoghurt or orange juice and their growth/survival monitored during enrichment at 37 °C or storage at 4 °C. The strains were also inoculated into minced beef and their thermal inactivation (D-values) examined at 55 °C, with and without a prior heat shock at 48 °C. The growth kin...

Genetic and antigenic relationship of foot-and-mouth disease virus serotype O isolates with the vaccine strain O1/BFS.

Science.gov (United States)

Xu, Wanhong; Zhang, Zhidong; Nfon, Charles; Yang, Ming

2018-05-15

Foot-and-mouth disease serotype O viruses (FMDV/O) are responsible for the most outbreaks in FMD endemic countries. O1/BFS is one of the recommended FMD/O vaccine strains by World Reference Laboratory for FMD. In the current study, FMDV/O1 BFS vaccine strain and serotype O field isolates (45) were analyzed phylogenetically and antigenically to gain more insight into the genetic and antigenic characteristics of the vaccine strain and field isolates. O1/BFS showed similarity with 89% of the field isolates using a virus neutralization test (VNT). The P1 region encoding the FMDV capsid was sequenced and analysed for 46 strains of FMDV/O. Phylogenetic analysis showed these viruses originated from five continents and covered eight of 11 reported topotypes. Five isolates that demonstrated low antigenic similarities with O1/BFS were analyzed for their antigenic variation at the known neutralizing antigenic sites. Three of the five isolates demonstrated unique amino acid substitutions at various antigenic sites. No unique amino acid substitutions were observed for the other two unmatched isolates. Positively selected residues were identified on the surface of the FMD virus capsid supporting that it is important to continuously monitor field isolates for their antigenic and phenotypic changes. In conclusion, the vaccine strain O1/BFS is likely to confer protection against 89% of the 45 FMDV/O isolates based on VNT. Thus O1/BFS vaccine strain is still suitable for use in global FMD serotype O outbreak control. Combining data from phylogenetic, molecular and antigenic analysis can provide improvements in the process of vaccine selection. Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.

Inhibition of Escherichia coli O157:H7 on stainless steel using Pseudomonas veronii biofilms.

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Kim, Y; Kim, H; Beuchat, L R; Ryu, J-H

2018-05-01

We produced a Pseudomonas veronii biofilm on the surface of a stainless steel that is inhibitory to Escherichia coli O157:H7. Pseudomonas veronii strain KACC 81051BP, isolated from lettuce, readily formed biofilm on the surface of stainless steel coupons (SSCs) immersed in tryptic soy broth at 25°C. Cells showed significantly (P ≤ 0·05) enhanced tolerance to desiccation stress (43% relative humidity (RH)) and retained antimicrobial activity against E. coli O157:H7. The number of E. coli O157:H7 (control; 4·1 ± 0·1 log CFU per coupon) on sterile SSCs decreased to 2·7 ± 0·2 log CFU per coupon after exposure to 43% RH at 25°C for 48 h, while the population of E. coli O157:H7 (4·1 ± 0·0 log CFU per coupon) on SSCs containing P. veronii biofilm decreased to below the theoretical detection limit (1·5 log CFU per coupon) within 24 h. The antimicrobial biofilm produced on stainless steel may have application in preventing cross-contamination by E. coli O157:H7 on other abiotic surfaces in food-contact environments. The presence of Escherichia coli O157:H7 on environmental surfaces of food manufacturing, transportation and storage facilities is a significant food safety concern because it can result in cross-contamination of food products. In this study, we developed a Pseudomonas veronii biofilm on the surface of a stainless steel that inhibits the growth of E. coli O157:H7. Since P. veronii in biofilm resists desiccation, it provides persistent antimicrobial activity. Information presented here provides novel and practical insights to developing biological strategies to inactivate E. coli O157:H7 on diverse surfaces in food processing and handling environments. © 2018 The Society for Applied Microbiology.

Molecular and Phenotypic Characterization of Escherichia coli O26:H8 among Diarrheagenic E. coli O26 Strains Isolated in Brazil

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Piazza, Roxane M. F.; Delannoy, Sabine; Fach, Patrick; Saridakis, Halha O.; Pedroso, Margareth Z.; Rocha, Letícia B.; Gomes, Tânia A. T.; Vieira, Mônica A. M.; Beutin, Lothar

2013-01-01

Escherichia coli strains of serogroup O26 comprise two distinct groups of pathogens, characterized as enteropathogenic E. coli (EPEC) and enterohemorrhagic E. coli (EHEC). Among the several genes related to type III secretion system-secreted effector proteins, espK was found to be highly specific for EHEC O26:H11 and its stx-negative derivative strains isolated in European countries. E. coli O26 strains isolated in Brazil from infant diarrhea, foods, and the environment have consistently been shown to lack stx genes and are thus considered atypical EPEC. However, no further information related to their genetic background is known. Therefore, in this study, we aimed to discriminate and characterize these Brazilian O26 stx-negative strains by phenotypic, genetic, and biochemical approaches. Among 44 isolates confirmed to be O26 isolates, most displayed flagellar antigen H11 or H32. Out of the 13 nonmotile isolates, 2 tested positive for fliCH11, and 11 were fliCH8 positive. The identification of genetic markers showed that several O26:H11 and all O26:H8 strains tested positive for espK and could therefore be discriminated as EHEC derivatives. The presence of H8 among EHEC O26 and its stx-negative derivative isolates is described for the first time. The interaction of three isolates with polarized Caco-2 cells and with intestinal biopsy specimen fragments ex vivo confirmed the ability of the O26 strains analyzed to cause attaching-and-effacing (A/E) lesions. The O26:H32 strains, isolated mostly from meat, were considered nonvirulent. Knowledge of the virulence content of stx-negative O26 isolates within the same serotype helped to avoid misclassification of isolates, which certainly has important implications for public health surveillance. PMID:23974139

Effect of heat treatment on the survival of Escherichia Coli O157:H7 ...

African Journals Online (AJOL)

The survival of Escherichia coli O157:H7 in raw milk treated in experimental pasteurizer was investigated in the year 2010. Raw milk was inoculated with different initial concentrations of E. coli O157:H7 and heated for 15 seconds at temperatures ranging from 69OC to 73OC. E. coli O157:H7 cells were not isolated from the ...

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

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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

Comparison of methods for the identification and sub-typing of O157 and non-O157 Escherichia coli serotypes and their integration into a polyphasic taxonomy approach

OpenAIRE

Prieto-Calvo M.A.; Omer M.K.; Alvseike O.; López M.; Alvarez-Ordóñez A.; Prieto M.

2016-01-01

Phenotypic, chemotaxonomic and genotypic data from 12 strains of Escherichia coli were collected, including carbon source utilisation profiles, ribotypes, sequencing data of the 16S–23S rRNA internal transcribed region (ITS) and Fourier transform-infrared (FT-IR) spectroscopic profiles. The objectives were to compare several identification systems for E. coli and to develop and test a polyphasic taxonomic approach using the four methodologies combined for the sub-typing of O157 and non-O157 E...

Assessments of Total and Viable Escherichia coli O157:H7 on Field and Laboratory Grown Lettuce

Science.gov (United States)

Moyne, Anne-Laure; Harris, Linda J.; Marco, Maria L.

2013-01-01

Leafy green produce has been associated with numerous outbreaks of foodborne illness caused by strains of Escherichia coli O157:H7. While the amounts of culturable E. coli O157:H7 rapidly decline after introduction onto lettuce in the field, it remains to be determined whether the reduction in cell numbers is due to losses in cell viability, cell injury and a subsequent inability to be detected by standard laboratory culturing methods, or a lack of adherence and hence rapid removal of the organism from the plants during application. To assess which of these options is most relevant for E. coli O157:H7 on leafy green produce, we developed and applied a propidium monoazide (PMA) real-time PCR assay to quantify viable (with PMA) and total (without PMA) E. coli O157:H7 cells on growth chamber and field-grown lettuce. E. coli O157:H7, suspended in 0.1% peptone, was inoculated onto 4-week-old lettuce plants at a level of approximately 106 CFU/plant. In the growth chamber at low relative humidity (30%), culturable amounts of the nontoxigenic E. coli O157:H7 strain ATCC 700728 and the virulent strain EC4045 declined 100 to 1000-fold in 24 h. Fewer E. coli O157:H7 cells survived when applied onto plants in droplets with a pipette compared with a fine spray inoculation. Total cells for both strains were equivalent to inoculum levels for 7 days after application, and viable cell quantities determined by PMA real-time PCR were approximately 104 greater than found by colony enumeration. Within 2 h after application onto plants in the field, the number of culturable E. coli ATCC 700728 was reduced by up to 1000-fold, whereas PCR-based assessments showed that total cell amounts were equivalent to inoculum levels. These findings show that shortly after inoculation onto plants, the majority of E. coli O157:H7 cells either die or are no longer culturable. PMID:23936235

Genetic diversity and antimicrobial resistance among isolates of Escherichia coli O157: H7 from feces and hides of super-shedders and low-shedding pen-mates in two commercial beef feedlots

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Stanford Kim

2012-09-01

Full Text Available Abstract Background Cattle shedding at least 104 CFU Escherichia coli O157:H7/g feces are described as super-shedders and have been shown to increase transmission of E. coli O157:H7 to other cattle in feedlots. This study investigated relationships among fecal isolates from super-shedders (n = 162, perineal hide swab isolates (PS from super-shedders (n = 137 and fecal isolates from low-shedder (4 CFU/g feces pen-mates (n = 496 using pulsed-field gel electrophoresis (PFGE. A subsample of these fecal isolates (n = 474 was tested for antimicrobial resistance. Isolates of E. coli O157:H7 were obtained from cattle in pens (avg. 181 head at 2 commercial feedlots in southern Alberta with each steer sampled at entry to the feedlot and prior to slaughter. Results Only 1 steer maintained super-shedder status at both samplings, although approximately 30% of super-shedders in sampling 1 had low-shedder status at sampling 2. A total of 85 restriction endonuclease digestion clusters (REPC; 90% or greater similarity and 86 unique isolates (P = 0.94. Only 2/21 super-shedders had fecal isolates in the same REPC at both samplings. Fecal and PS isolates from individual super-shedders generally belonged to different REPCs, although fecal isolates of E. coli O157:H7 from super- and low-shedders showed greater similarity (P P = 0.69, although all super-shedder isolates with antimicrobial resistance (n = 3 were resistant to multiple antimicrobials. Conclusions Super-shedders did not have increased antimicrobial resistance compared to low-shedder pen mates. Our data demonstrated that PFGE profiles of individual super-shedders varied over time and that only 1/162 steers remained a super-shedder at 2 samplings. In these two commercial feedlots, PFGE subtypes of E. coli O157:H7 from fecal isolates of super- and low-shedders were frequently different as were subtypes of fecal and perineal hide isolates from super-shedders.

Effect of Lactobacillus sp. isolates supernatant on Escherichia coli O157:H7 enhances the role of organic acids production as a factor for pathogen control

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Larissa B. Poppi

2015-04-01

Full Text Available Many attempts have been made to establish the control of foodborne pathogens through Lactobacillus isolates and their metabolism products with success being obtained in several situations. The aim of this study was to investigate the antagonistic effect of eight Lactobacillus isolates, including L. casei subsp. pseudoplantarum, L. plantarum, L. reuteri and L. delbrueckii subsp. delbrueckii, on the pathogenic Escherichia colistrain O157:H7. The inhibitory effect of pure cultures and two pooled cultures supernatants of Lactobacillus on the growth of pathogenic bacteria was evaluated by the spot agar method and by monitoring turbidity. Antimicrobial activity was confirmed for L. reuteri and L. delbrueckii subsp. delbrueckii and for a pool of lactic acid bacteria. The neutralized supernatant of the pool exerted a higher antimicrobial activity than that of the individual strains. Furthermore, D-lactic acid and acetic acid were produced during growth of the Lactobacillus isolates studied.

Complete Genome Sequence of an Escherichia coli O121:H19 Strain from an Outbreak in Canada Associated with Flour

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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

Prevalence and relatedness of Escherichia coli O157:H7 strains in the feces and on the hides and carcasses of U.S. meat goats at slaughter.

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Jacob, M E; Foster, D M; Rogers, A T; Balcomb, C C; Sanderson, M W

2013-07-01

We determined the prevalences of Escherichia coli O157:H7 in feces, hide, and carcasses of meat goats at a U.S. processing plant. Prevalences were 11.1%, 2.7%, and 2.7%, respectively. Sixteen pulsed-field gel electrophoresis (PFGE) subtypes were identified among 49 E. coli O157:H7 isolates, some of which were present on multiple sample types or collection days.

Prevalence and antibiogram of Escherichia coli O157 isolated from ...

African Journals Online (AJOL)

E. coli O157 from bovine carcass swabs and cecal contents were 9.3% and 7.3%, respectively. ..... Frequent use of tetracycline to treat animal dis- eases in ... Project entitled “Pneumonia, diarrhea and mastitis in food animals”. References.

In silico genomic analyses reveal three distinct lineages of Escherichia coli O157:H7, one of which is associated with hyper-virulence.

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Laing, Chad R; Buchanan, Cody; Taboada, Eduardo N; Zhang, Yongxiang; Karmali, Mohamed A; Thomas, James E; Gannon, Victor Pj

2009-06-29

Many approaches have been used to study the evolution, population structure and genetic diversity of Escherichia coli O157:H7; however, observations made with different genotyping systems are not easily relatable to each other. Three genetic lineages of E. coli O157:H7 designated I, II and I/II have been identified using octamer-based genome scanning and microarray comparative genomic hybridization (mCGH). Each lineage contains significant phenotypic differences, with lineage I strains being the most commonly associated with human infections. Similarly, a clade of hyper-virulent O157:H7 strains implicated in the 2006 spinach and lettuce outbreaks has been defined using single-nucleotide polymorphism (SNP) typing. In this study an in silico comparison of six different genotyping approaches was performed on 19 E. coli genome sequences from 17 O157:H7 strains and single O145:NM and K12 MG1655 strains to provide an overall picture of diversity of the E. coli O157:H7 population, and to compare genotyping methods for O157:H7 strains. In silico determination of lineage, Shiga-toxin bacteriophage integration site, comparative genomic fingerprint, mCGH profile, novel region distribution profile, SNP type and multi-locus variable number tandem repeat analysis type was performed and a supernetwork based on the combination of these methods was produced. This supernetwork showed three distinct clusters of strains that were O157:H7 lineage-specific, with the SNP-based hyper-virulent clade 8 synonymous with O157:H7 lineage I/II. Lineage I/II/clade 8 strains clustered closest on the supernetwork to E. coli K12 and E. coli O55:H7, O145:NM and sorbitol-fermenting O157 strains. The results of this study highlight the similarities in relationships derived from multi-locus genome sampling methods and suggest a "common genotyping language" may be devised for population genetics and epidemiological studies. Future genotyping methods should provide data that can be stored centrally and

In silico genomic analyses reveal three distinct lineages of Escherichia coli O157:H7, one of which is associated with hyper-virulence

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Karmali Mohamed A

2009-06-01

Full Text Available Abstract Background Many approaches have been used to study the evolution, population structure and genetic diversity of Escherichia coli O157:H7; however, observations made with different genotyping systems are not easily relatable to each other. Three genetic lineages of E. coli O157:H7 designated I, II and I/II have been identified using octamer-based genome scanning and microarray comparative genomic hybridization (mCGH. Each lineage contains significant phenotypic differences, with lineage I strains being the most commonly associated with human infections. Similarly, a clade of hyper-virulent O157:H7 strains implicated in the 2006 spinach and lettuce outbreaks has been defined using single-nucleotide polymorphism (SNP typing. In this study an in silico comparison of six different genotyping approaches was performed on 19 E. coli genome sequences from 17 O157:H7 strains and single O145:NM and K12 MG1655 strains to provide an overall picture of diversity of the E. coli O157:H7 population, and to compare genotyping methods for O157:H7 strains. Results In silico determination of lineage, Shiga-toxin bacteriophage integration site, comparative genomic fingerprint, mCGH profile, novel region distribution profile, SNP type and multi-locus variable number tandem repeat analysis type was performed and a supernetwork based on the combination of these methods was produced. This supernetwork showed three distinct clusters of strains that were O157:H7 lineage-specific, with the SNP-based hyper-virulent clade 8 synonymous with O157:H7 lineage I/II. Lineage I/II/clade 8 strains clustered closest on the supernetwork to E. coli K12 and E. coli O55:H7, O145:NM and sorbitol-fermenting O157 strains. Conclusion The results of this study highlight the similarities in relationships derived from multi-locus genome sampling methods and suggest a "common genotyping language" may be devised for population genetics and epidemiological studies. Future genotyping

Impact of persistent and nonpersistent generic Escherichia coli and Salmonella sp. recovered from a beef packing plant on biofilm formation by E. coli O157.

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Visvalingam, J; Ells, T C; Yang, X

2017-12-01

To examine the influence of meat plant Escherichia coli and Salmonella sp. isolates on E. coli O157 biofilm formation. Biofilm formation was quantified by crystal violet staining (A 570 nm ) and viable cell numbers for up to 6 days at 15°C. All five persistent E. coli genotypes formed strong biofilms when cultured alone or co-cultured with E. coli O157, with A 570 nm values reaching ≥4·8 at day 4, while only two of five nonpersistent genotypes formed such biofilms. For E. coli O157:H7 co-culture biofilms with E. coli genotypes 136 and 533, its numbers were ≥1·5 and ≥1 log CFU per peg lower than those observed for its mono-culture biofilm at days 2 and 4, respectively. The number of E. coli O157:NM in similar co-culture biofilms was 1 log CFU per peg lower than in its mono-culture biofilm at day 4 and 6, respectively. Salmonella sp. lowered the number of E. coli O157:NM by 0·5 log unit, once, at day 6. Generic E. coli may outcompete E. coli O157 strains while establishing biofilms. Findings advance knowledge regarding inter-strain competition for a similar ecological niche and may aid development of biocontrol strategies for E. coli O157 in food processing environments. © 2017 Her Majesty the Queen in Right of Canada. Journal of Applied Microbiology © 2017 The Society for Applied Microbiology Reproduced with the permission of the Minister of the Department of Agriculture and Agri-Food Canada.

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.

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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

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

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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.

Fatores de virulência em linhagens de Escherichia coli isoladas de mastite bovina Virulence factors in Escherichia coli strains isolated from bovine mastitis

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M.G. Ribeiro

2006-10-01

Full Text Available Avaliou-se a ocorrência de fatores de virulência e do sorotipo O157:H7 em 120 linhagens de Escherichia coli, isoladas de 80 casos de mastite clínica bovina e 40 de mastite subclínica. Verificou-se alfa-hemolisina em oito (6,7% linhagens, isoladas de cinco casos de mastite clínica e três de mastite subclínica e em nenhuma das estirpes detectou-se enteroemolisina. A presença de sideróforos foi encontrada em 11 (9,2% linhagens, sete de mastite clínica e quatro de subclínica. Em duas (1,7% estirpes isoladas de mastite subclínica, identificou-se enterotoxina STa. Observou-se efeito citopático em células vero compatível com a produção de verotoxina-VT em cinco (4,2% linhagens, duas de mastite clínica e três subclínicas. Em uma (0,8% linhagem isolada de mastite clínica, detectou-se efeito citopático compatível com o fator necrosante citotóxico. Nenhuma estirpe apresentou-se sorbitol-negativa no MacConkey-sorbitol, tampouco aglutinou com o sorotipo O157:H7. Os antimicrobianos mais efetivos foram polimixina B (97,5% e norfloxacina (95,8%. Observou-se multi-resistência a dois ou mais antimicrobianos em 24 (20% estirpes, principalmente com o uso de ampicilina e ceftiofur.The occurrence of different virulence factors and O157:H7 serotype investigation in 120 Escherichia coli strains isolated from clinical (80 cases and subclinical (40 cases bovine mastitis was evaluated. Alpha-haemolysin was detected in 8 (6.7% strains (5 clinical and 3 subclinical cases. None strain showed enterohaemolysin production. E. coli growth under iron restriction conditions (siderophores production was observed in 11 (9.2% strains (7 clinical and 4 subclinical cases. STa enterotoxin was detected in 2 (1.7% strains from subclinical cases. Cytotoxic effect in vero cells compatible with verotoxin-VT production was observed in 5 (4.2% strains (2 clinical and 3 subclinical cases. One strain (0.8% isolated from clinical mastitis showed cytophatic effect in vero

Frequency and risk-factors analysis of Escherichia coli O157:H7 in Bali-cattle.

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Suardana, I Wayan; Widiasih, Dyah Ayu; Nugroho, Widagdo Sri; Wibowo, Michael Haryadi; Suyasa, I Nyoman

2017-08-01

Cattle are known as the main reservoir of zoonotic agents verocytotoxin-producing Escherichia coli. These bacteria are usually isolated from calves with diarrhea and/or mucus and blood. Tolerance of these agents to the environmental conditions will strengthen of their transmission among livestock. A total of 238 cattle fecal samples from four sub-districts in Badung, Bali were used in this study. Epidemiological data observed include cattle age, sex, cattle rearing system, the source of drinking water, weather, altitude, and type of cage floor, the cleanliness of cage floor, the slope of cage floor, and the level of cattle cleanliness. The study was initiated by culturing of samples onto eosin methylene blue agar, then Gram stained, and tested for indole, methyl-red, voges proskauer, and citrate, Potential E.coli isolates were then cultured onto sorbitol MacConkey agar, and further tested using O157 latex agglutination test and H7 antisera. Molecular identification was performed by analysis of the 16S rRNA gene, and epidemiological data was analyzed using STATA 12.0 software. The results showed, the prevalence of E. coli O157:H7 in cattle at Badung regency was 6.30% (15/238) covering four sub districts i.e. Petang, Abiansemal, Mengwi, and Kuta which their prevalence was 8.62%(5/58), 10%(6/60), 3.33%(2/60), and 3.33(2/60)%, respectively. The analysis of 16S rRNA gene confirmed of isolates as an E. coli O157:H7 strain with 99% similarities. Furthermore, the risk factors analysis showed that the slope of the cage floor has a highly significant effect (P<0.05) to the distribution of infection. Consequently, implementing this factor must be concerned in order to decrease of infection. Copyright © 2017 Elsevier B.V. All rights reserved.

Prevalence and Antimicrobial Susceptibility Pattern of E. coli O157:H7 Isolated from Traditionally Marketed Raw Cow Milk in and around Asosa Town, Western Ethiopia.

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Disassa, Nigatu; Sibhat, Berhanu; Mengistu, Shimelis; Muktar, Yimer; Belina, Dinaol

2017-01-01

A cross-sectional study was conducted from October 2014 to July 2015 to determine the prevalence and populations of E. coli as well as the prevalence and antimicrobial susceptibility of E. coli O157:H7 isolated from raw milk. Biochemical and serological tests methods were used to confirm E. coli and E. coli O157:H7 and isolates were subjected to antimicrobial susceptibility test using the agar disc diffusion method. Out of 380 raw milk samples examined, 129 (33.9%) and 11 (2.9%) were contaminated with E. coli and E. coli O157:H7, respectively. The highest prevalence was recorded in samples obtained from vendors (39.1%, 4.978 ± 0.180 log 10 /ml) compared with samples from farmers (28.1%, 3.93 ± 0.01 log 10 /ml) with significant differences ( P = 0.02). The frequency of contamination was higher in the samples collected from milk that was stored and transported in plastic containers (39.4%) than in the containers made of stainless steel (23.0%) ( P = 0.002). The antimicrobial susceptibility profile showed that E. coli O157:H7 were resistant to tetracycline (81.8%), streptomycin (81.8%), and kanamycin (63.6%). Milk samples were produced and handled under poor hygienic conditions, stored, and transported in inappropriate containers and under temperature abuse conditions leading to high health risk to the consumers. Additional studies would be needed to establish association between the occurrences of E. coli O157:H7 in raw milk and all the risk factors involved in and around Asosa town.

Health Risk of Escherichia coli O157:H7 in Drinking Water and Meat and Meat Products and Vegetables to Diarrhoeic Confirmed and Non-Confirmed HIV/AIDS Patients

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Abong`O, B. O.; Momba, M. N. B.; Rodda, N.

The current study explored the health risk of E. coli O157:H7 to diarrhoeic confirmed and non-confirmed HIV/AIDS patients due to their exposure to presumed ingestion of water, meat products and vegetables ostensibly contaminated with E. coli O157:H7. Strains of E. coli O157:H7 were isolated by enrichment culture and on Cefixime-Telurite Sorbitol MacConkey agar. Average counts of presumptive E. coli O157 were used for dose-response assessment. Probability of infection to confirmed and non-confirmed HIV/AIDS patients was 20 and 27% from meat and meat products, 21% and 15% from vegetables and 100% due to ingestion of 1500 mL person-1 day-1 of water. Drinking water had higher probability of transmitting E. coli O157:H7 infections than meat and meat products and vegetables. Probability of E. coli O157:H7 infections were high for confirmed HIV/AIDS patients than for non-confirmed patients. Water and foods consumed by HIV/AIDS patients should be safe of any microbial contaminants, these waters and foods should as well be investigated for other enteric pathogens to establish their safety.

Diversity of Survival Patterns among Escherichia coli O157:H7 Genotypes Subjected to Food-Related Stress Conditions.

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Elhadidy, Mohamed; Álvarez-Ordóñez, Avelino

2016-01-01

The purpose of this study was to evaluate the resistance patterns to food-related stresses of Shiga toxin producing Escherichia coli O157:H7 strains belonging to specific genotypes. A total of 33 E. coli O157:H7 strains were exposed to seven different stress conditions acting as potential selective pressures affecting the transmission of E. coli O157:H7 to humans through the food chain. These stress conditions included cold, oxidative, osmotic, acid, heat, freeze-thaw, and starvation stresses. The genotypes used for comparison included lineage-specific polymorphism, Shiga-toxin-encoding bacteriophage insertion sites, clade type, tir (A255T) polymorphism, Shiga toxin 2 subtype, and antiterminator Q gene allele. Bacterial resistance to different stressors was calculated by determining D-values (times required for inactivation of 90% of the bacterial population), which were then subjected to univariate and multivariate analyses. In addition, a relative stress resistance value, integrating resistance values to all tested stressors, was calculated for each bacterial strain and allowed for a ranking-type classification of E. coli O157:H7 strains according to their environmental robustness. Lineage I/II strains were found to be significantly more resistant to acid, cold, and starvation stress than lineage II strains. Similarly, tir (255T) and clade 8 encoding strains were significantly more resistant to acid, heat, cold, and starvation stress than tir (255A) and non-clade 8 strains. Principal component analysis, which allows grouping of strains with similar stress survival characteristics, separated strains of lineage I and I/II from strains of lineage II, which in general showed reduced survival abilities. Results obtained suggest that lineage I/II, tir (255T), and clade 8 strains, which have been previously reported to be more frequently associated with human disease cases, have greater multiple stress resistance than strains of other genotypes. The results from this

Antibioticumresistentie in Escherichia coli O 157 geisoleerd tussen 1998 en 2003 in Nederland = Antibiotic resistance in Escherichia coli O157 isolated between 1998 and 2003 in The Netherlands

NARCIS (Netherlands)

Roest, H.I.J.; Liebana, E.; Wannet, W.; Duynhoven, van Y.; Veldman, K.T.; Mevius, D.J.

2007-01-01

Nederlands: Over het vóórkomen van antibioticumresistentie bij E. coli O157 in Nederland is weinig bekend. In deze studie werden tussen 1998 en 2003 218 humane en 247 niethumane isolaten onderzocht op antibioticumgevoeligheid. Het antibioticumresistentieniveau van E. coli O157 geïsoleerd uit de

Vaccination with killed whole-cells of Escherichia coli O157:H7 hha mutant emulsified with an adjuvant induced vaccine strain-specific serum antibodies and reduced E. coli O157:H7 fecal shedding in cattle

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Escherichia coli O157:H7 (O157) can cause from a mild diarrheal illness to hemorrhagic colitis and hemolytic uremic syndrome in humans. Cattle are the primary reservoir for O157 and fecal shedding of O157 by these animals is a major risk factor in contamination of cattle hides and carcasses at slaug...

Influence of Detection Methods in Characterizing Escherichia coli O157:H7 in Raw Goat Meat Using Conventional and Molecular Methods.

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Tabashsum, Zajeba; Nazneen, Mafruha; Ahsan, C R; Bari, M L; Yasmin, M

2016-01-01

　Presence of Escherichia coli O157:H7 on fresh goat meat samples (n= 40) of Dhaka city was analyzed using conventional and molecular methods. A total of 86 presumptive E. coli O157:H7 colonies were isolated from 60% of the samples using selective agar plating method. After conventional biochemical assay followed by API 20E assay, only 11 isolates were found to be E. coli O157:H7. Further serological test identified only four isolates that has strong agglutination reaction against anti-H7 sensitized latex. The biochemically and serologically confirmed isolates were then screened for major virulence factors include eaeA, rfbE, fliC, stx1 and stx2 genes by PCR. PCR analysis of positive isolates showed, 10 isolates were eaeA and rfbE genes positive but fliC gene was only in six, indicating that these isolates were H7 positive with flagellum antigens which might not expressed or detected in serotyping tests. Multiplex PCR against eaeA, stx1 and stx2 genes of the isolates showed similar results as when done individually. These results revealed that only 7% of the primary presumptive E. coli O157:H7 was found to be stx producing E. coli O157:H7 and thus greatly influenced the detection of the pathogen in meat samples.

The antimicrobial peptide cathelicidin protects mice from Escherichia coli O157:H7-mediated disease.

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Milan Chromek

Full Text Available This study investigated the role of the antimicrobial peptide cathelicidin in Escherichia coli O157:H7 infection and subsequent renal damage. Mouse and human cathelicidin, CRAMP and LL-37, respectively, killed E. coli O157:H7 in vitro. Intestines from healthy wild-type (129/SvJ and cathelicidin-knock-out (Camp(-/- mice were investigated, showing that cathelicidin-deficient mice had a thinner colonic mucus layer compared with wild-type mice. Wild-type (n = 11 and cathelicidin-knock-out (n = 11 mice were inoculated with E. coli O157:H7. Cathelicidin-deficient animals exhibited higher fecal counts of E. coli O157:H7 and bacteria penetrated the mucus forming attaching-and-effacing lesions to a much higher extent than in wild-type animals. Cathelicidin knock-out mice developed symptoms (9/11 as well as anemia, thrombocytopenia and extensive renal tubular damage while all cathelicidin-producing mice remained asymptomatic with normal laboratory findings. When injected with Shiga toxin intraperitoneally, both murine strains developed the same degree of renal tubular damage and clinical disease indicating that differences in sensitivity to infection between the murine strains were related to the initial intestinal response. In conclusion, cathelicidin substantially influenced the antimicrobial barrier in the mouse colon mucosa. Cathelicidin deficiency lead to increased susceptibility to E. coli O157:H7 infection and subsequent renal damage. Administration of cathelicidin or stimulation of endogenous production may prove to be novel treatments for E. coli O157:H7-induced hemolytic uremic syndrome.

British Escherichia coli O157 in Cattle Study (BECS): to determine the prevalence of E. coli O157 in herds with cattle destined for the food chain.

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Henry, M K; Tongue, S C; Evans, J; Webster, C; McKENDRICK, I J; Morgan, M; Willett, A; Reeves, A; Humphry, R W; Gally, D L; Gunn, G J; Chase-Topping, M E

2017-11-01

Escherichia coli O157 are zoonotic bacteria for which cattle are an important reservoir. Prevalence estimates for E. coli O157 in British cattle for human consumption are over 10 years old. A new baseline is needed to inform current human health risk. The British E. coli O157 in Cattle Study (BECS) ran between September 2014 and November 2015 on 270 farms across Scotland and England & Wales. This is the first study to be conducted contemporaneously across Great Britain, thus enabling comparison between Scotland and England & Wales. Herd-level prevalence estimates for E. coli O157 did not differ significantly for Scotland (0·236, 95% CI 0·166-0·325) and England & Wales (0·213, 95% CI 0·156-0·283) (P = 0·65). The majority of isolates were verocytotoxin positive. A higher proportion of samples from Scotland were in the super-shedder category, though there was no difference between the surveys in the likelihood of a positive farm having at least one super-shedder sample. E. coli O157 continues to be common in British beef cattle, reaffirming public health policy that contact with cattle and their environments is a potential infection source.

Persistence of Escherichia coli O157:H7 in dairy fermentation systems.

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Dineen, S S; Takeuchi, K; Soudah, J E; Boor, K J

1998-12-01

We examined (i) the persistence of Escherichia coli O157:H7 as a postpasteurization contaminant in fermented dairy products; (ii) the ability of E. coli O157:H7 strains with and without the general stress regulatory protein, RpoS, to compete with commercial starter cultures in fermentation systems; and (iii) the survival of E. coli O157:H7 in the yogurt production process. In commercial products inoculated with 10(3) CFU/ml, E. coli O157:H7 was recovered for up to 12 days in yogurt (pH 4.0), 28 days in sour cream (pH 4.3), and at levels > 10(2) CFU/ml at 35 days in buttermilk (pH 4.1). For the starter culture competition trials, the relative inhibition of E. coli O157:H7 in the experimental fermentation systems was, in decreasing order, thermophilic culture mixture, Lactobacillus delbrueckii subsp. bulgaricus R110 alone, Lactococcus lactis subsp. lactis D280 alone, Lactococcus lactis subsp. cremoris D62 alone, and Streptococcus thermophilus C90 alone showing the least inhibition. Recovery of the rpoS mutant was lower than recovery of its wild-type parent by 72 h or earlier in the presence of individual starter cultures. No E. coli O157:H7 were recovered after the curd formation step in yogurt manufactured with milk inoculated with 10(5) CFU/ml. Our results show that (i) postprocessing entry of E. coli O157:H7 into fermented dairy products represents a potential health hazard; (ii) commercial starter cultures differ in their ability to reduce E. coli O157:H7 CFU numbers in fermentation systems; and (iii) the RpoS protein appears to most effectively contribute to bacterial survival in the presence of conditions that are moderately lethal to the cell.

Detection of Escherichia Coli O157:H7 in Fecal Samples in Meat Goats

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Mobley, Ray; Madden, Uford; Brooks-Walter, Alexis

2004-01-01

Studies have reported the isolation of Escherichia coli (E. coli)O157:H7 from pork, lamb and poultry products, and from other animals including deer, horses, dogs, birds and humans. There is limited or no information on the presence of the organism in goats. The objectives of this study were to determine if E. coli O157:H7 was naturally occurring…

Clonal Diversity of Chilean Isolates of Enterohemorrhagic Escherichia coli from Patients with Hemolytic-Uremic Syndrome, Asymptomatic Subjects, Animal Reservoirs, and Food Products

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Rios, Maritza; Prado, Valeria; Trucksis, Michele; Arellano, Carolina; Borie, Consuelo; Alexandre, Marcela; Fica, Alberto; Levine, Myron M.

1999-01-01

To determine clonal relationship among Chilean enterohemorrhagic Escherichia coli (EHEC) strains from different sources (clinical infections, animal reservoirs, and food), 54 EHEC isolates (44 of E. coli O157, 5 of E. coli O111, and 5 of E. coli O26) were characterized for virulence genes by colony blot hybridization and by pulsed-field gel electrophoresis (PFGE). By colony blotting, 12 different genotypes were identified among the 44 E. coli O157 isolates analyzed, of which the genetic profile stx1+ stx2+ hly+ eae+ was the most prevalent. All human O157 strains that were associated with sporadic cases of hemolytic-uremic syndrome (HUS) carried both the stx1 and stx2 toxin-encoding genes and were eaeA positive. Only 9 of 13 isolates from human controls were stx1+ stx2+, and 8 carried the eaeA gene. Comparison of profiles obtained by PFGE of XbaI-digested genomic DNA showed a great diversity among the E. coli O157 isolates, with 37 different profiles among 39 isolates analyzed. Cluster analysis of PFGE profiles showed a wide distribution of clinical isolates obtained from HUS cases and asymptomatic individuals and a clonal relationship among O157 isolates obtained from HUS cases and pigs. Analysis of virulence genes showed that a correlation exists among strains with the genotype stx1+ stx2+ eae+ and pathogenic potential. A larger difference in the PFGE restriction patterns was observed among the EHEC strains of serogroups O26 and O111. These results indicate that several different EHEC clones circulate in Chile and suggest that pigs are an important animal reservoir for human infections by EHEC. Guidelines have been proposed for better practices in the slaughter of animals in Chile. PMID:9986852

Escherichia coli O157:H7 Outbreak Associated with Restaurant Beef Grinding.

Science.gov (United States)

Torso, Lauren M; Voorhees, Ronald E; Forest, Stephen A; Gordon, Andrew Z; Silvestri, Sharon A; Kissler, Bonnie; Schlackman, Jessica; Sandt, Carol H; Toma, Paul; Bachert, Joel; Mertz, Kristen J; Harrison, Lee H

2015-07-01

Escherichia coli O157:H7 is a common cause of foodborne illness in the United States. Beef ground at establishments regulated by the U.S. Department of Agriculture, Food Safety and Inspection Service is routinely tested for E. coli O157:H7. Prior to December 2013, boxed beef product (wholesale cuts of beef, such as beef loin, packaged into bags and boxed for shipping) was not always tested for this pathogen. Downstream processors or retailers may grind the product; and, if the ground beef is not cooked to the recommended temperature, pathogens on the exterior of the beef introduced to the interior through grinding may survive. On 18 October 2013, the Allegheny County Health Department identified two E. coli O157:H7 cases, both of whom were food handlers at restaurant A, a restaurant that ground locally produced boxed beef for hamburgers on site. Case finding was conducted through public messaging, employee surveys, and disease surveillance. All potential cases were interviewed using a standard questionnaire. A confirmed case was defined as laboratory-confirmed E. coli O157:H7 with exposure to restaurant A. A probable case was defined as a patient with compatible symptoms and exposure to restaurant A but without laboratory confirmation. All human and food isolates were characterized by pulsed-field gel electrophoresis and multilocus variable-number tandem repeat analysis. The analysis identified 14 confirmed and 10 probable cases of E. coli; 18 nonintact ground beef samples tested positive for E. coli O157:H7. Nine confirmed cases were restaurant A employees. All confirmed cases recalled eating a restaurant A hamburger in the 10 days before illness onset; most cases reported consuming medium to rare hamburgers. Multiple pulsed-field gel electrophoresis and multilocus variable-number tandem repeat analysis patterns were identified among both the human and ground beef isolates, and the patient isolates matched those found in ground beef samples. Restaurant A

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.

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.

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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.

FimH has a strain and host-cell type dependent role in adherence of E. coli O157:H7 super-shedder strains to host cells

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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...

The Escherichia coli O157:H7 bovine rumen fluid proteome reflects adaptive bacterial responses.

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Kudva, Indira T; Stanton, Thaddeus B; Lippolis, John D

2014-02-21

To obtain insights into Escherichia coli O157:H7 (O157) survival mechanisms in the bovine rumen, we defined the growth characteristics and proteome of O157 cultured in rumen fluid (RF; pH 6.0-7.2 and low volatile fatty acid content) obtained from rumen-fistulated cattle fed low protein content "maintenance diet" under diverse in vitro conditions. Bottom-up proteomics (LC-MS/MS) of whole cell-lysates of O157 cultured under anaerobic conditions in filter-sterilized RF (fRF; devoid of normal ruminal microbiota) and nutrient-depleted and filtered RF (dRF) resulted in an anaerobic O157 fRF-and dRF-proteome comprising 35 proteins functionally associated with cell structure, motility, transport, metabolism and regulation, but interestingly, not with O157 virulence. Shotgun proteomics-based analysis using isobaric tags for relative and absolute quantitation used to further study differential protein expression in unfiltered RF (uRF; RF containing normal rumen microbial flora) complemented these results. Our results indicate that in the rumen, the first anatomical compartment encountered by this human pathogen within the cattle gastrointestinal tract (GIT), O157 initiates a program of specific gene expression that enables it to adapt to the in vivo environment, and successfully transit to its colonization sites in the bovine GIT. Further experiments in vitro using uRF from animals fed different diets and with additional O157 strains, and in vivo using rumen-fistulated cattle will provide a comprehensive understanding of the adaptive mechanisms involved, and help direct evolution of novel modalities for blocking O157 infection of cattle.

Synergistic interaction in dual-species biofilms formation by Escherichia coli O157:H7 and Ralstonia spp

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Introduction: Ralstonia spp., a heterotrophic bacterium that are isolated from produce processing environments as part of the native microflora, have strong potentials for formaing biofilms on various surfaces. When co-cultured, Escherichia coli O157:H7 (EcO157) and Ralstonia spp. displayed a synerg...

Determination of the Thermal Inactivation Kinetics of Listeria monocytogenes, Salmonella enterica, and Escherichia coli O157:H7 and non-O157 in Buffer and a Spinach Homogenate.

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Monu, Emefa Angelica; Valladares, Malcond; D'Souza, Doris H; Davidson, P Michael

2015-08-01

Produce has been associated with a rising number of foodborne illness outbreaks. While much produce is consumed raw, some is treated with mild heat, such as blanching or cooking. The objectives of this research were to compare the thermal inactivation kinetics of Listeria monocytogenes, Salmonella enterica, Shiga toxin-producing Escherichia coli (STEC) O157:H7, and non-O157 STEC in phosphate-buffered saline (PBS; pH 7.2) and a spinach homogenate and to provide an estimate of the safety of mild heat processes for spinach. Five individual strains of S. enterica, L. monocytogenes, STEC O157:H7, and non-O157 STEC were tested in PBS in 2-ml glass vials, and cocktails of the organisms were tested in blended spinach in vacuum-sealed bags. For Listeria and Salmonella at 56 to 60°C, D-values in PBS ranged from 4.42 ± 0.94 to 0.35 ± 0.03 min and 2.11 ± 0.14 to 0.16 ± 0.03 min, respectively. D-values at 54 to 58°C were 5.18 ± 0.21 to 0.53 ± 0.04 min for STEC O157:H7 and 5.01 ± 0.60 to 0.60 ± 0.13 min for non-O157 STEC. In spinach at 56 to 60°C, Listeria D-values were 11.77 ± 2.18 to 1.22 ± 0.12 min and Salmonella D-values were 3.51 ± 0.06 to 0.47 ± 0.06 min. D-values for STEC O157:H7 and non-O157 STEC were 7.21 ± 0.17 to 1.07 ± 0.11 min and 5.57 ± 0.38 to 0.99 ± 0.07 min, respectively, at 56 to 60°C. In spinach, z-values were 4.07 ± 0.16, 4.59 ± 0.26, 4.80 ± 0.92, and 5.22 ± 0.20°C for Listeria, Salmonella, STEC O157:H7, and non-O157 STEC, respectively. Results indicated that a mild thermal treatment of blended spinach at 70°C for less than 1 min would result in a 6-log reduction of all pathogens tested. These findings may assist the food industry in the design of suitable mild thermal processes to ensure food safety.

Survival and interaction of Escherichia coli O104:H4 on Arabidopsis thaliana and lettuce (Lactuca sativa) in comparison to E. coli O157:H7: Influence of plant defense response and bacterial capsular polysaccharide.

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Jang, Hyein; Matthews, Karl R

2018-06-01

Shiga toxin-producing Escherichia coli (STEC) has been associated with illnesses and outbreaks linked to fresh vegetables, prompting a growing public health concern. Most studies regarding interactions of STEC on fresh produce focused on E. coli O157:H7. Limited information is available about survival or fitness of E. coli O104:H4, non-O157 pathogen that was linked to one of the largest outbreaks of hemolytic uremic syndrome in 2011. In this study, survival of E. coli O104:H4 was evaluated on Arabidopsis thaliana plant and lettuce for 5 days compared with E. coli O157:H7, and expression of pathogenesis-realted gene (PR1; induction of plant defense response) was examined by reverse transcription quantitative PCR, and potential influence of capsular polysaccharide (CPS) on the bacterial fitness on plant was investigated. Populations of E. coli O104:H4 strains (RG1, C3493, and LpfA) on Arabidopsis and lettuce were significantly (P E. coli O157:H7 strains (7386 and sakai) at day 5 post-inoculation, indicating E. coli O104:H4 may have better survival ability on the plants. In addition, the E. coli O104:H4 strains produced significantly (P E. coli O157:H7 strains. RG1 strain (1.5-fold) initiated significantly (P E. coli O157:H7 strains 7386 (2.9-fold) and sakai (2.7-fold). Collectively, the results in this study suggests that different level of CPS production and plant defense response initiated by each STEC strain might influence the bacterial survival or persistence on plants. The present study provides better understanding of survival behavior of STEC, particularly E. coli O104:H4, using a model plant and vegetable under pre-harvest conditions with plant defense response. Copyright © 2018 Elsevier Ltd. All rights reserved.

Acid and alcohol tolerance of Escherichia coli O157:H7 in pulque, a typical Mexican beverage.

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Gómez-Aldapa, Carlos A; Díaz-Cruz, Claudio A; Villarruel-López, Angelica; Del Refugio Torres-Vitela, M; Rangel-Vargas, Esmeralda; Castro-Rosas, Javier

2012-03-01

Pulque is a traditional Mexican fermented alcoholic beverage produced from the nectar of maguey agave plants. No data exist on the behavior of Escherichia coli O157:H7 in agave nectar and pulque. An initial trial was done of the behavior of E. coli O157:H7 during fermentation of nectar from a single producer, a nectar mixture from different producers and "seed" pulque. A second trial simulating artisanal pulque production was done by contaminating fresh nectar with a cocktail of three E. coli O157:H7 strains, storing at 16 ° and 22 °C for 14 h, adding seed pulque and fermenting until pulque was formed. A third trial used pulque from the second trial stored at 22 °C as seed to ferment fresh nectar at 22 °C for 48 h (fermentation cycle). This procedure was repeated for an additional two fermentation cycles. During incubation at 16 ° or 22 °C in the first trial, the E. coli O157:H7 strains multiplied in both the single producer nectar and nectar mixture, reaching maximum concentration at 12h. E. coli O157:H7 cell concentration then decreased slowly, although it survived at least 72 h in both fermented nectars. E. coli O157:H7 did not multiply in the seed pulque but did survive at least 72 h. In the second trial, the numbers of E. coli O157:H7 increased approximately 1.5 log CFU/ml at 22 °C and 1.2 log CFU/ml at 16 °C after 14 h. After seed pulque was added, E. coli O157:H7 concentration decreased to approximately 2 log CFU/ml, and then remained constant until pulque was produced. In the third trial, the E. coli O157:H7 cells multiplied and survived during at least three nectar fermentation cycles. The results suggest that E. coli O157:H7 can develop acid and alcohol tolerance in pulque, and constitutes a public health risk for pulque consumers. Copyright © 2011 Elsevier B.V. All rights reserved.

Identification of E.coli O157:H7 in Intestinal and Urinary Tract Infection in Samawah City .

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Mouna Akeel Hamed Al-Oebady

2017-02-01

Full Text Available This study was conducted to isolate E.coli O157: H7 as an important zoonotic pathogen from 150 samples (75 bloody stools and 75 urine samples of patients at many age groups range from one to 50 years old and for both sexes were collected from patients suffering from diarrhea and urinary tract infection who attend the Samawah Teaching Hospital for pediatrics and Gynecology of AL-Muthanna Governorate. The results revealed that 120 out of 150 were positive to E.coli O157:H7 at a percentage (80%. The number of E. coli isolates in bloody stool were 67(89.3% and urine samples were 53(70.6% gave positive results to E.coli O157:H7 .

Does enterohemorrhagic Escherichia coli O157:H7 enter the viable but nonculturable state in salted salmon roe?

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Makino, S I; Kii, T; Asakura, H; Shirahata, T; Ikeda, T; Takeshi, K; Itoh, K

2000-12-01

An outbreak caused by salted salmon roe contaminated with enterohemorrhagic Escherichia coli O157 occurred in Japan in 1998. Since about 0.75 to 1.5 viable cells were estimated to cause infection, we presumed that O157 might enter the viable but nonculturable (VNC) state in salted salmon roe and consequently that viable cell numbers might be underestimated. Although patient-originating O157 cells could not grow on agar plates after 72 h of incubation in 13% NaCl, they were resuscitated in yeast extract broth, and more than 90% of the cells were shown to be viable by fluorescent staining, suggesting that almost all of them could enter the VNC state in NaCl water. Roe-originating O157 was resistant to NaCl because it could grow on agar after 72 h of incubation in NaCl water, but about 20% of cells appeared to enter the VNC state. Therefore, germfree mice were infected with O157 to examine the resuscitation of cells in the VNC state and the retention of pathogenicity. O157 that originated in roe, but not patients, killed mice and was isolated from the intestine. However, these isolates had become sensitive to NaCl. O157 cells of roe origin incubated in normal media also killed mice and were isolated from the intestine, but they also became transiently NaCl sensitive. We therefore propose that bacterial cells might enter the VNC state under conditions of stress, such as those encountered in vivo or in high salt concentrations, and then revive when those conditions have eased. If so, the VNC state in food is potentially dangerous from a public health viewpoint and may have to be considered at the time of food inspection. Finally, the establishment of a simple recovery system for VNC cells should be established.

Dynamic of Mutational Events in Variable Number Tandem Repeats of Escherichia coli O157:H7

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A. V. Bustamante

2013-01-01

Full Text Available VNTRs regions have been successfully used for bacterial subtyping; however, the hypervariability in VNTR loci is problematic when trying to predict the relationships among isolates. Since few studies have examined the mutation rate of these markers, our aim was to estimate mutation rates of VNTRs specific for verotoxigenic E. coli O157:H7. The knowledge of VNTR mutational rates and the factors affecting them would make MLVA more effective for epidemiological or microbial forensic investigations. For this purpose, we analyzed nine loci performing parallel, serial passage experiments (PSPEs on 9 O157:H7 strains. The combined 9 PSPE population rates for the 8 mutating loci ranged from 4.4 × 10−05 to 1.8 × 10−03 mutations/generation, and the combined 8-loci mutation rate was of 2.5 × 10−03 mutations/generation. Mutations involved complete repeat units, with only one point mutation detected. A similar proportion between single and multiple repeat changes was detected. Of the 56 repeat mutations, 59% were insertions and 41% were deletions, and 72% of the mutation events corresponded to O157-10 locus. For alleles with up to 13 UR, a constant and low mutation rate was observed; meanwhile longer alleles were associated with higher and variable mutation rates. Our results are useful to interpret data from microevolution and population epidemiology studies and particularly point out that the inclusion or not of O157-10 locus or, alternatively, a differential weighting data according to the mutation rates of loci must be evaluated in relation with the objectives of the proposed study.

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.

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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.

Detección de Escherichia coli O157: H7 en carne picada fresca y hamburguesas congeladas Escherichia coli O157: H7 detection in fresh ground beef and hamburgers

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M. A. Marzocca

2006-03-01

points of our supermarket chain, totalling 37 and 43, respectively. These samples were processed using the EC selective enrichment broth containing novobiocin, then followed by the application of an immunocapture method (TECRA E. COLI O157 IMMUNOCAPTURE TM ECOICM 20, and later isolation in MacConkey sorbitol agar with cefixime and potassium tellurite, in a chromogenic medium. The suspected strains were genotypically characterized by PCR detection of the stx1, stx2, eaeA, and EHEC-hlyA genes, and by a colony blot hybridization assay. Serotyping, antimicrobial susceptibility patterns, and production of Stx by a specific citotoxicity assay on Vero cells were also determined. E coli O157:H7 was isolated in only one fresh ground beef sample (2,7%, identified as gene eae (+/stx2/EHEC-hlyA.

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

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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,

Curli fimbriae are conditionally required in Escherichia coli O157:H7 for initial attachment and biofilm formation.

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Carter, Michelle Qiu; Louie, Jacqueline W; Feng, Doris; Zhong, Wayne; Brandl, Maria T

2016-08-01

Several species of enteric pathogens produce curli fimbriae, which may affect their interaction with surfaces and other microbes in nonhost environments. Here we used two Escherichia coli O157:H7 outbreak strains with distinct genotypes to understand the role of curli in surface attachment and biofilm formation in several systems relevant to fresh produce production and processing. Curli significantly enhanced the initial attachment of E. coli O157:H7 to spinach leaves and stainless steel surfaces by 5-fold. Curli was also required for E. coli O157:H7 biofilm formation on stainless steel and enhanced biofilm production on glass by 19-27 fold in LB no-salt broth. However, this contribution was not observed when cells were grown in sterile spinach lysates. Furthermore, both strains of E. coli O157:H7 produced minimal biofilms on polypropylene in LB no-salt broth but considerable amounts in spinach lysates. Under the latter conditions, curli appeared to slightly increase biofilm production. Importantly, curli played an essential role in the formation of mixed biofilm by E. coli O157:H7 and plant-associated microorganisms in spinach leaf washes, as revealed by confocal microscopy. Little or no E. coli O157:H7 biofilms were detected at 4 °C, supporting the importance of temperature control in postharvest and produce processing environments. Published by Elsevier Ltd.

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

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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

Application of whole genome sequence data in analyzing the molecular epidemiology of Shiga toxin-producing Escherichia coli O157:H7/H.

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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

Biocontrol of Escherichia coli O157:H7 on fresh-cut spinach and lettuce using a bacteriophage cocktail

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The effect of an E. coli O157:H7-specific bacteriophage cocktail (EcoShield™) was evaluated against nalidixic acid resistant (NalR) E. coli O157:H7 strains in either a) laboratory medium or b) on leafy greens. Laboratory medium cultures were inoculated with 5 log CFU/ml and treated with 7 log PFU/ml...

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.

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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.

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.

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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.

Impact of dry chilling on the genetic diversity of Escherichia coli on beef carcasses and on the survival of E. coli and E. coli O157.

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Visvalingam, Jeyachchandran; Liu, Yang; Yang, Xianqin

2017-03-06

The objective of this study was to examine the effect of dry chilling on the genetic diversity of naturally occurring Escherichia coli on beef carcasses, and to examine whether two populations of E. coli recovered from carcasses during chilling and E. coli O157 differed in their response to desiccation. Isolates of E. coli were obtained from beef carcasses during a 67h dry chilling process and were genotyped using multiple-locus variable-number tandem-repeat analysis (MLVA). Ten E. coli genotypes found only at 0h (group A) and found more than once (group B), as well as five strains of E. coli O157 (group C) were inoculated on stainless steel coupons and their survival was examined after exposure to 75 and 100% relative humidity (RH) at 0 or 35°C for 67h. A total of 450 E. coli isolates were obtained, with 254, 49, 49, 51, 23, 20, and 4 from 0, 1, 2, 4, 6, 8 and 24h of chilling, respectively. No E. coli were recovered at 67h. MLVA of the isolates revealed 173 distinct genotypes. Genetic diversity of E. coli isolates, defined as ratio of the number of isolates to the number of genotypes, remained between 2.3 and 1.3 during the 24h of chilling. All strains inoculated on stainless steel coupons and exposed to 75% RH at 35°C were completely inactivated, irrespective of their groups. Inactivation of E. coli of the three groups was not significantly (P>0.05) different by exposure to 75% RH at 0°C. The findings indicate that the genetic diversity of E. coli on beef carcasses was not affected by dry chilling. In addition, inactivation of E. coli genotypes and E. coli O157 by desiccation on stainless steel simulating dry chilling conditions did not differ significantly (P>0.05). Thus, dry chilling may be used as an effective antimicrobial intervention for beef carcasses. Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.

Detection of Escherichia coli O157:H7 and Salmonella in ground beef by a bead-free quantum dot-facilitated isolation method.

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Wang, Luxin; Wu, Chung-Shieh; Fan, Xudong; Mustapha, Azlin

2012-05-01

The aims of this study were to introduce a new immunological bead-free cell detection method using quantum dots (QDs) as reporter markers for foodborne pathogen detection. QDs are nanosized particles with long-term photostability, high quantum yield, broad absorption spectra, and narrow, symmetric emission and high signal-to-noise ratio. The chemical compound [(1-ethyl-3-3-dimethylaminopropyl) carbodiimide hydrochloride] (EDC) and protein A were used as crosslinkers for manufacturing QD-antibody conjugates. To minimize the inhibition of QD fluorescence by the magnetic beads, the beads were removed after the primary pathogen isolation and before fluorescence measurement. Detection signals were increased four-fold after employing the bead-free isolation method. With a 24-h enrichment, the bead-free QD-facilitated detection method was able to detect 10 CFU/g Escherichia coli O157:H7 and Salmonella from artificially contaminated ground beef. To our knowledge, this detection method is the first research that combined a new EDC-protein A QD-labeling technique and bead-free fluorescence measurement to detect E. coli O157:H7 and Salmonella in ground beef. Copyright © 2012 Elsevier B.V. All rights reserved.

E. coli O157 on Scottish cattle farms: Evidence of local spread and persistence using repeat cross-sectional data

Science.gov (United States)

2014-01-01

Background Escherichia coli (E. coli) O157 is a virulent zoonotic strain of enterohaemorrhagic E. coli. In Scotland (1998-2008) the annual reported rate of human infection is 4.4 per 100,000 population which is consistently higher than other regions of the UK and abroad. Cattle are the primary reservoir. Thus understanding infection dynamics in cattle is paramount to reducing human infections. A large database was created for farms sampled in two cross-sectional surveys carried out in Scotland (1998 - 2004). A statistical model was generated to identify risk factors for the presence of E. coli O157 on farms. Specific hypotheses were tested regarding the presence of E. coli O157 on local farms and the farms previous status. Pulsed-field gel electrophoresis (PFGE) profiles were further examined to ascertain whether local spread or persistence of strains could be inferred. Results The presence of an E. coli O157 positive local farm (average distance: 5.96km) in the Highlands, North East and South West, farm size and the number of cattle moved onto the farm 8 weeks prior to sampling were significant risk factors for the presence of E. coli O157 on farms. Previous status of a farm was not a significant predictor of current status (p = 0.398). Farms within the same sampling cluster were significantly more likely to be the same PFGE type (p Scottish E. coli O157 paves the way for future research into the mechanisms of transmission which should help with the design of control measures to reduce E. coli O157 from livestock-related sources. PMID:24766709

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 ...

The first pathogenic Yersinia enterocolitica bioserotype 4/O:3 strain isolated from a hunted wild boar (Sus scrofa) in Poland.

Science.gov (United States)

Bancerz-Kisiel, A; Platt-Samoraj, A; Szczerba-Turek, A; Syczyło, K; Szweda, W

2015-10-01

The objective of this study was to identify the bioserotypes and virulence markers of Yersinia enterocolitica strains isolated from wild boars in Poland. Bacteriological examination of 302 rectal swabs from 151 wild boars resulted in the isolation of 40 Y. enterocolitica strains. The majority of the examined strains (n = 30), belonged to bioserotype 1A/NI. The presence of individual Y. enterocolitica strains belonging to bioserotypes 1B/NI (3), 1A/O:8 (2), 1A/O:27 (2), 2/NI (1), 2/O:9 (1) and 4/O:3 (1) was also demonstrated. Amplicons corresponding to ail and ystA genes were observed only in one Y. enterocolitica strain--bioserotype 4/O:3. The ail and ystB gene amplicons were noted in 11 Y. enterocolitica biotype 1A strains, although single amplicons of ystB gene were found in 28 of the tested samples. In four out of eight cases when two Y. enterocolitica strains were isolated from the same animal, the strains differed in biotype, serotype or virulence markers. The European population of wild boars continues to grow and spread to new areas, therefore, wild boars harbouring potentially pathogenic Y. enterocolitica 4/O:3 strains pose a challenge to public health.

Escherichia coli O157:H7 - An Emerging Pathogen in foods of Animal Origin

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Ch. Bindu Kiranmayi

Full Text Available Escherichia coli O157:H7 is an emerging public health concern in most countries of the world. E. coli O157:H7 was known to be a human pathogen for nearly 24 years. EHEC O157 infection is estimated to be the fourth most costly food borne disease in Canada and USA, not counting the cost of possible litigation. E. coli O157:H7 and Salmonella are the leading causes of produce related outbreaks, accounting for 20 and 30% respectively. The authority of the Federal Meat Inspection Act, FSIS (Food Safety and Inspection Service declared Escherichia coli O157:H7, an adulterant in raw ground beef and enforced “zero tolerance” (USDA-FSIS, 17 December 1998. Because of the severity of these illnesses and the apparent low infective dose (less than 10 cells, Escherichia coli O157:H7 is considered one of the most serious of known food borne pathogens. Escherichia coli O157:H7 is mainly pathogenic to human but in cattle and other animals, it did not induce any clinical disease except diarrhea. So, these animals act as carriers to Escherichia coli O157:H7. The majority transmission is through eating of undercooked contaminated ground meat and consumption of raw milk, raw vegetables, fruits contaminated by water, cheese, curd and also through consumption of sprouts, lettuce and juice. The conventional isolation procedure includes growth in enrichment broth like modified EC (E. coli broth or modified tryptic soy broth (mTSB Since the infection primarily occurs via faeco-oral route, the preventive measures include food hygiene measures like proper cooking of meat, consumption of pasteurized milk, washing fruits and vegetables especially those to be eaten raw and drinking chlorine treated water and personnel hygiene measures like washing hands after toilet visits. [Veterinary World 2010; 3(8.000: 382-389

Efficacy of integrated treatment of UV light and low dose gamma irradiation on Escherichia coli O157:H7 and Salmonella enterica on grape tomatoes

Science.gov (United States)

Efficacy of integrated treatment of UVC and low dose Gamma irradiation to inactivate mixed Strains of Escherichia coli O157:H7 and Salmonella enterica inoculated on whole Grape tomatoes was evaluated. A mixed bacterial cocktail composed of a three strain mixture of E. coli O157:H7 (C9490, E02128 an...

Effect of Neem (Azadirachta indica on the Survival of Escherichia coli O157:H7 in Dairy Manure

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Subbarao V. Ravva

2015-07-01

Full Text Available Escherichia coli O157:H7 (EcO157 shed in cattle manure can survive for extended periods of time and intervention strategies to control this pathogen at the source are critical as produce crops are often grown in proximity to animal raising operations. This study evaluated whether neem (Azadirachta indica, known for its antimicrobial and insecticidal properties, can be used to amend manure to control EcO157. The influence of neem materials (leaf, bark, and oil on the survival of an apple juice outbreak strain of EcO157 in dairy manure was monitored. Neem leaf and bark supplements eliminated the pathogen in less than 10 d with a D-value (days for 90% elimination of 1.3 d. In contrast, nearly 4 log CFU EcO157/g remained after 10 d in neem-free manure control. The ethyl acetate extractable fraction of neem leaves was inhibitory to the growth of EcO157 in LB broth. Azadirachtin, a neem product with insect antifeedant properties, failed to inhibit EcO157. Application of inexpensive neem supplements to control pathogens in manure and possibly in produce fields may be an option for controlling the transfer of foodborne pathogens from farm to fork.

Topological data analysis of Escherichia coli O157:H7 and non-O157 survival in soils

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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 ...

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

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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.

Cross contamination of Escherichia coli O157:H7 between lettuce and wash water during home-scale washing.

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Jensen, Dane A; Friedrich, Loretta M; Harris, Linda J; Danyluk, Michelle D; Schaffner, Donald W

2015-04-01

Lettuce and leafy greens have been implicated in multiple foodborne disease outbreaks. This study quantifies cross contamination between lettuce pieces in a small-scale home environment. A five-strain cocktail of relevant Escherichia coli O157:H7 strains was used. Bacterial transfer between single inoculated lettuce leaf pieces to 10 non-inoculated lettuce leaf pieces that were washed in a stainless steel bowl of water for 30 s, 1 min, 2 min, and 5 min was quantified. Regardless of washing time, the wash water became contaminated with 90-99% of bacteria originally present on the inoculated lettuce leaf piece. The E. coli O157:H7 concentration on initially inoculated leaf pieces was reduced ∼ 2 log CFU. Each initially uncontaminated lettuce leaf piece had ∼ 1% of the E. coli O157:H7 from the inoculated lettuce piece transferred to it after washing, with more transfer occurring during the shortest (30 s) and longest (5 min) wash times. In all cases the log percent transfer rates were essentially normally distributed. In all scenarios, most of the E. coli O157:H7 (90-99%) transferred from the inoculated lettuce pieces to the wash water. Washing with plain tap water reduces levels of E. coli O157:H7 on the inoculated lettuce leaf pieces, but also spreads contamination to previously uncontaminated leaf pieces. Copyright © 2014 Elsevier Ltd. All rights reserved.

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

Escherichia coli O157:H7- patógeno alimentar emergente / Escherichia coli O157:H7 - emerging food pathogen

Directory of Open Access Journals (Sweden)

Cheila Minéia Daniel de Paula

2014-11-01

has gained great prominence in the last 20 years, due to the severity of its outbreaks. Until recently, Brazil was considered to be free of this pathogen, but the scientific literature and epidemiological records prove otherwise. This article thus aims to perform an integrative review of the literature, focusing on the characteristics, isolation, and detection methods, and the epidemiological data of E. coli O157: H7 in Brazil and the world.

Escherichia coli O157:H7 and rectoanal junction cell interactome

Science.gov (United States)

Introduction. Cattle are the primary E. coli O157 (O157) reservoir and principal source of human infection. The anatomical site of O157 persistence is the bovine recto-anal (RAJ) junction; hence, an in-depth understanding of O157-RAJ interactions will help develop novel modalities to limit O157 in c...

An outbreak of Vero cytotoxin producing Escherichia coli O157 infection associated with takeaway sandwiches.

LENUS (Irish Health Repository)

McDonnell, R J

1997-12-12

An outbreak of food poisoning due to Escherichia coli O157 phage type 2 Vero cytotoxin 2 affected 26 people in southern counties of England in May and June 1995. The organism was isolated from faecal specimens from 23 patients, 16 of whom lived in Dorset and seven in Hampshire. Isolates were indistinguishable by phage typing, Vero cytotoxin gene typing, restriction fragment length polymorphism, and pulsed field gel electrophoresis. Three associated cases, linked epidemiologically to the outbreak, were confirmed serologically by detection of antibodies to E. coli O157 lipopolysaccharide. Twenty-two of the 26 patients were adults: four were admitted to hospital with haemorrhagic colitis. Four cases were children: two were admitted to hospital with haemolytic uraemic syndrome (HUS). There were no deaths. Although E. coli O157 was not isolated from any food samples, illness was associated with having eaten cold meats in sandwiches bought from two sandwich producers, in Weymouth and in Portsmouth. Both shops were supplied by the same wholesaler, who kept no records and obtained cooked meats from several sources in packs that did not carry adequate identification marks. It was, therefore, impossible to trace back to the original producer or to investigate further to determine the origin of contamination with E. coli O157. To protect the public health it is essential that all wholesale packs of ready-to-eat food carry date codes and the producer\\'s identification mark. Detailed record keeping should be part of hazard analysis critical control point (HACCP) systems and should be maintained throughout the chain of distribution from the producer to retail outlets.

Isolation and molecular characterization of Salmonella enterica, Escherichia coli O157:H7 and Shigella spp. from meat and dairy products in Egypt.

Science.gov (United States)

Ahmed, Ashraf M; Shimamoto, Tadashi

2014-01-03

Foodborne pathogens are a major threat to food safety, especially in developing countries where hygiene and sanitation facilities are often poor. Salmonella enterica, Escherichia coli O157:H7 and Shigella spp. are among the major causes of outbreaks of foodborne diseases. This large-scale study investigated the prevalence of these foodborne pathogens in meat (beef and chicken) and dairy products collected from street vendors, butchers, retail markets and slaughterhouses in Egypt. A total of 1600 food samples (800 meat products and 800 dairy products) were analyzed using culture and PCR based methods. S. enterica, E. coli O157:H7 and Shigella spp. were detected in 69 (4.3%), 54 (3.4%) and 27 (1.7%) samples respectively. S. enterica serovar Typhimurium, S. enterica serovar Enteritidis, S. enterica serovar Infantis and non-typable serovars were detected in 28 (1.8%), 22 (1.4%), 16 (1.0%) and 3 (0.1%) samples respectively. All E. coli O157:H7 isolates were positive for stx1 and/or stx2 virulence toxin genes. Shigella flexneri, Shigella sonnei and Shigella dysenteriae were detected in 18 (1.2%), 7 (0.4%) and 2 (0.1%) samples respectively. The incidences of S. enterica and Shigella spp. were higher in meat products (53; 6.6% and 16; 2.0%, respectively) than in dairy products (16; 2.0% and 11; 1.4%, respectively), while, E. coli O157:H7 was higher in dairy products (29; 3.6%) than in meat products (25; 3.1%). The incidence of foodborne pathogens in meat and dairy products was determined in a large-scale survey in Africa. © 2013.

Distribution and content of class 1 integrons in different Vibrio cholerae O-serotype strains isolated in Thailand

DEFF Research Database (Denmark)

Dalsgaard, Anders; Forslund, Anita; Serichantalergs, Oralak

2000-01-01

only a single antibiotic resistance gene. Although resistance genes in class 1 integrons were found in strains from the same epidemic, as well as in unrelated non-O1, non-O139 strains isolated from children with diarrhea, they were found to encode only some of the antibiotic resistance expressed...

Rapid and selective detection of E. coli O157:H7 combining phagomagnetic separation with enzymatic colorimetry.

Science.gov (United States)

Zhang, Yun; Yan, Chenghui; Yang, Hang; Yu, Junping; Wei, Hongping

2017-11-01

Mammal IgG antibodies are normally used in conventional immunoassays for E. coli O157:H7, which could lead to false positive results from the presence of protein A producing S. aureus. In this study, a natural specific bacteriophage was isolated and then conjugated with magnetic beads as a capture element in a sandwich format for the rapid and selective detection of E. coli O157:H7. To the best of our knowledge, it was the first time to utilize a natural bacteriophage to develop a phagomagnetic separation combined with colorimetric assay for E. coli O157:H7. The method has an overall time less than 2h with a detection limit of 4.9×10 4 CFU/mL. No interference from S. aureus was observed. Furthermore, the proposed method was successfully applied to detect E. coli O157:H7 in spiked skim milk. The proposed detection system provided a potential method for E. coli O157:H7 and other pathogenic bacteria in food samples. Copyright © 2017 Elsevier Ltd. All rights reserved.

Genotypic Characterization of Escherichia coli O157:H7 Isolates from Different Sources in the North-West Province, South Africa, Using Enterobacterial Repetitive Intergenic Consensus PCR Analysis

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Collins Njie Ateba

2014-05-01

Full Text Available In many developing countries, proper hygiene is not strictly implemented when animals are slaughtered and meat products become contaminated. Contaminated meat may contain Escherichia coli (E. coli O157:H7 that could cause diseases in humans if these food products are consumed undercooked. In the present study, a total of 94 confirmed E. coli O157:H7 isolates were subjected to the enterobacterial repetitive intergenic consensus (ERIC polymerase chain reaction (PCR typing to generate genetic fingerprints. The ERIC fragments were resolved by electrophoresis on 2% (w/v agarose gels. The presence, absence and intensity of band data were obtained, exported to Microsoft Excel (Microsoft Office 2003 and used to generate a data matrix. The unweighted pair group method with arithmetic mean (UPGMA and complete linkage algorithms were used to analyze the percentage of similarity and matrix data. Relationships between the various profiles and/or lanes were expressed as dendrograms. Data from groups of related lanes were compiled and reported on cluster tables. ERIC fragments ranged from one to 15 per isolate, and their sizes varied from 0.25 to 0.771 kb. A large proportion of the isolates produced an ERIC banding pattern with three duplets ranging in sizes from 0.408 to 0.628 kb. Eight major clusters (I–VIII were identified. Overall, the remarkable similarities (72% to 91% between the ERIC profiles for the isolate from animal species and their corresponding food products indicated some form of contamination, which may not exclude those at the level of the abattoirs. These results reveal that ERIC PCR analysis can be reliable in comparing the genetic profiles of E. coli O157:H7 from different sources in the North-West Province of South Africa.

Genotypic characterization of Escherichia coli O157:H7 isolates from different sources in the North-West Province, South Africa, using enterobacterial repetitive intergenic consensus PCR analysis.

Science.gov (United States)

Ateba, Collins Njie; Mbewe, Moses

2014-05-30

In many developing countries, proper hygiene is not strictly implemented when animals are slaughtered and meat products become contaminated. Contaminated meat may contain Escherichia coli (E. coli) O157:H7 that could cause diseases in humans if these food products are consumed undercooked. In the present study, a total of 94 confirmed E. coli O157:H7 isolates were subjected to the enterobacterial repetitive intergenic consensus (ERIC) polymerase chain reaction (PCR) typing to generate genetic fingerprints. The ERIC fragments were resolved by electrophoresis on 2% (w/v) agarose gels. The presence, absence and intensity of band data were obtained, exported to Microsoft Excel (Microsoft Office 2003) and used to generate a data matrix. The unweighted pair group method with arithmetic mean (UPGMA) and complete linkage algorithms were used to analyze the percentage of similarity and matrix data. Relationships between the various profiles and/or lanes were expressed as dendrograms. Data from groups of related lanes were compiled and reported on cluster tables. ERIC fragments ranged from one to 15 per isolate, and their sizes varied from 0.25 to 0.771 kb. A large proportion of the isolates produced an ERIC banding pattern with three duplets ranging in sizes from 0.408 to 0.628 kb. Eight major clusters (I-VIII) were identified. Overall, the remarkable similarities (72% to 91%) between the ERIC profiles for the isolate from animal species and their corresponding food products indicated some form of contamination, which may not exclude those at the level of the abattoirs. These results reveal that ERIC PCR analysis can be reliable in comparing the genetic profiles of E. coli O157:H7 from different sources in the North-West Province of South Africa.

Pathogenic Potential to Humans of Bovine Escherichia coli O26, Scotland

Science.gov (United States)

Rosser, Tracy; Allison, Lesley J.; Courcier, Emily; Evans, Judith; McKendrick, Iain J.; Pearce, Michael C.; Handel, Ian; Caprioli, Alfredo; Karch, Helge; Hanson, Mary F.; Pollock, Kevin G.J.; Locking, Mary E.; Woolhouse, Mark E.J.; Matthews, Louise; Low, J. Chris; Gally, David L.

2012-01-01

Escherichia coli O26 and O157 have similar overall prevalences in cattle in Scotland, but in humans, Shiga toxin–producing E. coli O26 infections are fewer and clinically less severe than E. coli O157 infections. To investigate this discrepancy, we genotyped E. coli O26 isolates from cattle and humans in Scotland and continental Europe. The genetic background of some strains from Scotland was closely related to that of strains causing severe infections in Europe. Nonmetric multidimensional scaling found an association between hemolytic uremic syndrome (HUS) and multilocus sequence type 21 strains and confirmed the role of stx2 in severe human disease. Although the prevalences of E. coli O26 and O157 on cattle farms in Scotland are equivalent, prevalence of more virulent strains is low, reducing human infection risk. However, new data on E. coli O26–associated HUS in humans highlight the need for surveillance of non-O157 enterohemorrhagic E. coli and for understanding stx2 phage acquisition. PMID:22377426

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

Occurrence of verotoxigenic E.coli in cow feces and antimicrobial resistance of the isolates in cattle farms in Shahrekord area

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Mojtaba Bonyadian

2017-09-01

Full Text Available Introduction:Escherichia coli is a common bacterium in the intestinal microflora of warm-blooded animals. They are routinely shed into the environment through feces and can contaminate water and soil, and, consequently fruits and vegetables .Enterohemorrhagic E. coli strains are recently emerged group of food-borne pathogens that are a significant public health threat. This group causes bloody diarrhea and hemolytic uremic syndrome (HUS, and the disease is prevalent in developed countries. The purpose of this study was to isolate and identify the E.coli O157: H7 and other verotoxigenic ones and major virulence genes (rfbE, eaeA, stx1, stx2 in fecal swab samples by PCR in Shahrekord area. Materials and methods: In Spring and Summer 2015, 400 cow fecal swab samples were collected from farms in Shahrekord area. Bacteriological and biochemical examinations were done for detection of E.coli. PCR assay was done for identification of O157:H7 serotype and other verotoxigenic E. coli using rfbE, eae, stx1 and stx2 genes. Results: E. coli O157:H7 was not detected in any strains tested. But PCR showed that out of 384 E.coli strain, 104(27/08% isolates carried stx1 gene, 36(9/37% carried stx2 gene and 16 (4.16% carried both stx1 and stx2 genes. Intimin (eaeA gene was detected in 280(72/91% of the isolates. Among verotoxigenic strain antibiotic resistance to Tetracycline 87/1%, Ampiciline 51/62%, Cefotaxime 48/38%, Gentamycin 25/81%,, Ciprofeloxacin 3/22% and Sulfamethoxazol 3/22% were observed. Discussion and conclusion: According to the results, although the serotype O157: H7 did not isolate from the feces of cattle but other verotoxigenic strains that showed high resistance  to antibiotic were isolated so it is a risk for human health.

Survival of Escherichia coli O157:H7 in needle-tenderized dry cured Westphalian ham.

Science.gov (United States)

Graumann, Gary H; Holley, Richard A

2007-09-15

Westphalian ham is a dry cured, ready-to-eat product that is manufactured without a lethal heat treatment. Hams are preserved by a process that involves curing, fermenting, smoking and drying, which may take 3 months or more to complete. The process can be accelerated by tenderizing the meat with solid needles, to increase the rate of cure-salt diffusion throughout muscle tissues. In this study, intact hams were immersed in a solution containing a five strain cocktail of Escherichia coli O157:H7 at 8 log cfu/mL, to determine whether needle treatment before cure application would internalize organisms from the surface. In two trials, the survival of E. coli O157:H7 on external surfaces and within deep tissues after needle treatment was followed during the ripening of hams. The injured E. coli O157:H7 cells were recovered by plating samples on pre-poured Tryptic Soy Agar plates which were incubated for 3 to 4 h at 35 degrees C, overlaid with Sorbitol MacConkey Agar containing cefixime and tellurite and re-incubated at 35 degrees C for 48 to 72 h. Inoculated-injected hams initially carried E. coli O157:H7 at numbers of 7.3 and 4.6 log cfu/g E. coli O157:H7 on the surface and inside, respectively. After 112 d of ripening, which included 79 d of drying, no E. coli O157:H7 were detected at the surface of hams following enrichment, whereas in deep tissue the organism was recovered at numbers of 3.1 log cfu/g. The Westphalian ham ripening procedure evidently was not adequate to eliminate E. coli O157:H7 internalized by needle tenderization.

Whole Genome Sequencing for Genomics-Guided Investigations of Escherichia coli O157:H7 Outbreaks.

Science.gov (United States)

Rusconi, Brigida; Sanjar, Fatemeh; Koenig, Sara S K; Mammel, Mark K; Tarr, Phillip I; Eppinger, Mark

2016-01-01

Multi isolate whole genome sequencing (WGS) and typing for outbreak investigations has become a reality in the post-genomics era. We applied this technology to strains from Escherichia coli O157:H7 outbreaks. These include isolates from seven North America outbreaks, as well as multiple isolates from the same patient and from different infected individuals in the same household. Customized high-resolution bioinformatics sequence typing strategies were developed to assess the core genome and mobilome plasticity. Sequence typing was performed using an in-house single nucleotide polymorphism (SNP) discovery and validation pipeline. Discriminatory power becomes of particular importance for the investigation of isolates from outbreaks in which macrogenomic techniques such as pulse-field gel electrophoresis or multiple locus variable number tandem repeat analysis do not differentiate closely related organisms. We also characterized differences in the phage inventory, allowing us to identify plasticity among outbreak strains that is not detectable at the core genome level. Our comprehensive analysis of the mobilome identified multiple plasmids that have not previously been associated with this lineage. Applied phylogenomics approaches provide strong molecular evidence for exceptionally little heterogeneity of strains within outbreaks and demonstrate the value of intra-cluster comparisons, rather than basing the analysis on archetypal reference strains. Next generation sequencing and whole genome typing strategies provide the technological foundation for genomic epidemiology outbreak investigation utilizing its significantly higher sample throughput, cost efficiency, and phylogenetic relatedness accuracy. These phylogenomics approaches have major public health relevance in translating information from the sequence-based survey to support timely and informed countermeasures. Polymorphisms identified in this work offer robust phylogenetic signals that index both short- and

Escherichia coli O157:H7 Converts Plant-Derived Choline to Glycine Betaine for Osmoprotection during Pre- and Post-harvest Colonization of Injured Lettuce Leaves

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Russell A. Scott

2017-12-01

Full Text Available Plant injury is inherent to the production and processing of fruit and vegetables. The opportunistic colonization of damaged plant tissue by human enteric pathogens may contribute to the occurrence of outbreaks of foodborne illness linked to produce. Escherichia coli O157:H7 (EcO157 responds to physicochemical stresses in cut lettuce and lettuce lysates by upregulation of several stress response pathways. We investigated the tolerance of EcO157 to osmotic stress imposed by the leakage of osmolytes from injured lettuce leaf tissue. LC-MS analysis of bacterial osmoprotectants in lettuce leaf lysates and wound washes indicated an abundant natural pool of choline, but sparse quantities of glycine betaine and proline. Glycine betaine was a more effective osmoprotectant than choline in EcO157 under osmotic stress conditions in vitro. An EcO157 mutant with a deletion of the betTIBA genes, which are required for biosynthesis of glycine betaine from imported choline, achieved population sizes twofold lower than those of the parental strain (P < 0.05 over the first hour of colonization of cut lettuce in modified atmosphere packaging (MAP. The cell concentrations of the betTIBA mutant also were 12-fold lower than those of the parental strain (P < 0.01 when grown in hypertonic lettuce lysate, indicating that lettuce leaf cellular contents provide choline for osmoprotection of EcO157. To demonstrate the utilization of available choline by EcO157 for osmoadaptation in injured leaf tissue, deuterated (D-9 choline was introduced to wound sites in MAP lettuce; LC-MS analysis revealed the conversion of D9-choline to D-9 glycine betaine in the parental strain, but no significant amounts were observed in the betTIBA mutant. The EcO157 ΔbetTIBA-ΔotsBA double mutant, which is additionally deficient in de novo synthesis of the compatible solute trehalose, was significantly less fit than the parental strain after their co-inoculation onto injured lettuce leaves and

In vitro safety assessments and antimicrobial activities of Lactobacillus rhamnosus strains isolated from a fermented mare's milk.

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Aryantini, Ni Putu Desy; Yamasaki, Eiki; Kurazono, Hisao; Sujaya, I Nengah; Urashima, Tadasu; Fukuda, Kenji

2017-03-01

Safety and probiotic characteristics such as antimicrobial activities of three Lactobacillus rhamnosus strains, FSMM15, FSMM22 and FSMM26, previously isolated as potential probiotics from fermented mare's milk were investigated. The three FSMM strains were susceptible to ampicillin, gentamycin, kanamycin, streptomycin, tetracycline and chloramphenicol, whereas they were resistant to erythromycin (minimal inhibitory concentration (MIC) = 4-8 µg/mL) and clindamycin (MIC = 4 µg/mL); bioconversion of bile salts, hemolytic activity and mucin degradation activity were negative; enzymatic activities of α-chymotrypsin and β-glucosidase were detected, but those of α-galactosidase, β-glucuronidase and N-acetyl-β-glucosaminidase, were undetectable. Among the strains, strain FSMM15 was chosen as a safer probiotic candidate due mainly to the lack of plasminogen binding ability. Despite lower acid production of strain FSMM15 than others, its cell-free culture supernatant inhibited growths of Salmonella Typhimurium LT-2, Shigella sonnei, Listeria monocytogenes, and Escherichia coli O157 with comparable levels of ampicillin, suggesting a favorable aspect of strain FSMM15 as a probiotic strain. © 2016 Japanese Society of Animal Science.

Screening for virulence genes in Escherichia coli O157:H7 obtained ...

African Journals Online (AJOL)

Eighty (80) sources of drinking water comprising boreholes (24), streams (3), wells (29), pipe-borne (5) and 19 sachet water samples were collected between March 2014 and February 2015. Escherichia coli (E.coli) O157:H7 was isolated by enrichment in Tryptone soy broth at elevated temperature and streaking on Eosin ...

Synergistic effect of enterocin AS-48 in combination with outer membrane permeabilizing treatments against Escherichia coli O157:H7.

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Ananou, S; Gálvez, A; Martínez-Bueno, M; Maqueda, M; Valdivia, E

2005-01-01

To determine the effects of outer membrane (OM) permeabilizing agents on the antimicrobial activity of enterocin AS-48 against Escherichia coli O157:H7 CECT 4783 strain in buffer and apple juice. We determined the influence of pH, EDTA, sodium tripolyphosphate (STPP) and heat on E. coli O157:H7 CECT 4783 sensitivity to enterocin AS-48 in buffer and in apple juice. Enterocin AS-48 was not active against intact cells of E. coli O157:H7 CECT 4783 at neutral pH. However, cells sublethally injured by OM permeabilizing agents (EDTA, STPP, pH 5, pH 8.6 and heat) became sensitive to AS-48, decreasing the amount of bacteriocin required for inhibition of E. coli O157:H7 CECT 4783. The results presented indicate that enterocin AS-48 could potentially be applied with a considerably wider range of protective agents, such as OM permeabilizing agents, with increased efficacy in inhibiting E. coli O157:H7. Results from this study support the potential use of enterocin AS-48 to control E. coli O157:H7 in combination with other hurdles.

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

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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.

Effect of severe weather events on the shedding of Shiga toxigenic Escherichia coli in slaughter cattle and phenotype of serogroup O157 isolates.

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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.

Identification and Prevalence of Escherichia coli and Escherichia coli O157: H7 in Foods

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Ancuta Mihaela Rotar

2013-11-01

Full Text Available The objective of this study is to investigate the incidence of Escherichia coli in animal and non-animal foods, and mainly the incidence of the serotype O157: H7 producing verotoxin. The presence of common Escherichia coli and Escherichia coli O157: H7 in various foods (of animal and non animal origin was performed in Transylvania area. We analyzed a total of one hundred forty-one samples of minced meat, one hundred twenty-six samples of meat , twenty six samples of meat products, five samples of alcoholic beverages, three samples of seafood, one hundred samples of cheese from pasteurized milk, seventeen samples of butter, four samples of vegetables and one sample of milk powder, using the standard cultural method and Vidas Eco method for E. coli O157: H7 strains. E. coli was identified in 50 samples of minced meat, 55 samples of meat prepared, 4 samples of meat products, 2 samples of alcoholic beverages, 25 samples of cheese from pasteurized milk, 6 samples of butter and 1 sample of vegetables. In this study were not been identified any foods contaminated with the E. coli O157: H7 serotype. The results of this reasearch have demostrated that E. coli wich represents a hygienic indicator of recent food contamination, can be destroyed with heat treatment and hygienic handling of foods. Our country over the years has been among the few countries where the incidence of the E. coli O157: H7 serotype has been minimal.

Predictors and risk factors for the intestinal shedding of Escherichia coli O157 among working donkeys (Equus asinus) in Nigeria

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Jedial, Jesse T.; Shittu, Aminu; Tambuwal, Faruk M.; Abubakar, Mikail B.; Garba, Muhammed K.; Kwaga, Jacob P.; Fasina, Folorunso O.

2015-01-01

Objectives Escherichia coli are an important group of bacteria in the normal gastrointestinal system but can sometimes cause infections in domestic animals and man. Donkeys are routinely used as multipurpose animal but details of burdens of potentially infectious bacteria associated with it are limited. The prevalence and associations between intestinal shedding of E. coli O157 and animal characteristics and management factors were studied among 240 randomly selected working donkeys in north-western Nigeria. Design Four local government areas, of Sokoto State in north-western Nigeria were recruited in this study. A multistage randomised cluster design was used to select subjects and donkey owners within selected zones. Confirmation of infection was based on bacterial culture, isolation and biochemical test for E. coli O157 from faecal samples. Results Of the total bacteria isolated, 203 of the 329 (61.70 per cent) were E. coli, 76 of which was E. coli serotype O157. A multivariable logistic regression model was used to examine the relation between intestinal shedding of E. coli O157 and selected variables. The analysis yielded five potential predictors of shedding: soft faeces in donkeys, Akaza and Fari ecotypes of donkey were positive predictors while maize straw as feed and sampling during the cold dry period were negative predictors. Conclusions This study concludes that controlling intestinal shedding of E. coli O157 among working donkeys in Nigeria is possible using the identified predictors in planning appropriate interventions to reduced human risk of infection. PMID:26392892

Culture supernatants from V. cholerae O1 El Tor strains isolated from different geographic areas induce cell vacuolation and cytotoxicity.

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Vidal, Jorge E; Enríquez-Rincón, Fernando; Giono-Cerezo, Silvia; Ribas-Aparicio, Rosa María; Figueroa-Arredondo, Paula

2009-01-01

To investigate whether the HlyA-induced vacuolating effect is produced by V. cholerae O1 ElTor strains isolated from different geographic origins, including Mexico. Supernatant-induced haemolysis, vacuolating activity and cytotoxicity in Vero cells were recorded. PCR, RFLP analysis and molecular cloning were performed. All ElTor strains analyzed induced cellular vacuolation. Ribotype 2 strains isolates from the U.S. gulf coast yielded the highest titer of vacuolating activity. Eight of nine strains were haemolytic, while all strains were PCR positive for the hlyA gene. We cloned the hlyA gene from two ElTor strains, a toxigenic (2514-88, ctxAB+) and a non-toxigenic Mexican strain (CM 91-3, ctxAB-). Supernatant from those recombinant E. coli strains induced haemolysis, cell vacuolation and cytotoxicity. RFLP-PCR analysis revealed similarities in the hlyA gene from all strains tested. The HlyA-induced vacuolating effect is a widespread phenotype of epidemic V. cholerae O1 ElTor strains.

Thermal tolerance of acid-adapted and unadapted Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes in cantaloupe juice and watermelon juice.

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Sharma, M; Adler, B B; Harrison, M D; Beuchat, L R

2005-01-01

A study was performed to determine D values of acid-adapted and unadapted cells of Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes in cantaloupe juice and watermelon juice. Salmonella enterica serotype Poona, S. enterica serotype Saphra, two strains of E. coli O157:H7, and two strains of L. monocytogenes were grown in tryptic soy broth (TSB) and TSB supplemented with 1% glucose for 24 h at 37 degrees C. Decimal reduction times (D values) of cells suspended in unpasteurized cantaloupe juice and watermelon juice were determined. Acid-adapted cells of Salmonella and E. coli O157:H7, but not L. monocytogenes, had increased thermal tolerance compared with cells that were not acid-adapted. There was no correlation between soluble solids content of the two types of juice and thermal resistance. Growth of Salmonella and E. coli O157:H7 in cantaloupe juice, watermelon juice, or other acidic milieu, either in preharvest or postharvest environments, may result in cross protection to heat. The pasteurization conditions necessary to achieve elimination of pathogens from these juices would consequently have to be more severe if cells are habituated to acidic environments. Insights from this study provide guidance to developing pasteurization processes to eliminate Salmonella, E. coli O157:H7, and L. monocytogenes in cantaloupe juice and watermelon juice.

A novel approach to investigate the uptake and internalization of Escherichia coli O157:H7 in spinach cultivated in soil and hydroponic media

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Internalization of E. coli O157:H7 into spinach plants through root uptake is a potential route of contamination. A Tn7-based plasmid vector was used to insert the green fluorescent protein (gfp) gene into the attTn7 site in the E. coli chromosome. Three gfp-labeled E. coli inocula, O157:H7 strains ...

Contaminated Stream Water as Source for Escherichia coli O157 Illness in Children.

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Probert, William S; Miller, Glen M; Ledin, Katya E

2017-07-01

In May 2016, an outbreak of Shiga toxin-producing Escherichia coli O157 infections occurred among children who had played in a stream flowing through a park. Analysis of E. coli isolates from the patients, stream water, and deer and coyote scat showed that feces from deer were the most likely source of contamination.

Comparison of methods for the identification and sub-typing of O157 and non-O157 Escherichia coli serotypes and their integration into a polyphasic taxonomy approach

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Prieto-Calvo M.A.

2016-12-01

Full Text Available Phenotypic, chemotaxonomic and genotypic data from 12 strains of Escherichia coli were collected, including carbon source utilisation profiles, ribotypes, sequencing data of the 16S–23S rRNA internal transcribed region (ITS and Fourier transform-infrared (FT-IR spectroscopic profiles. The objectives were to compare several identification systems for E. coli and to develop and test a polyphasic taxonomic approach using the four methodologies combined for the sub-typing of O157 and non-O157 E. coli. The nucleotide sequences of the 16S–23S rRNA ITS regions were amplified by polymerase chain reaction (PCR, sequenced and compared with reference data available at the GenBank database using the Basic Local Alignment Search Tool (BLAST . Additional information comprising the utilisation of carbon sources, riboprint profiles and FT-IR spectra was also collected. The capacity of the methods for the identification and typing of E. coli to species and subspecies levels was evaluated. Data were transformed and integrated to present polyphasic hierarchical clusters and relationships. The study reports the use of an integrated scheme comprising phenotypic, chemotaxonomic and genotypic information (carbon source profile, sequencing of the 16S–23S rRNA ITS, ribotyping and FT-IR spectroscopy for a more precise characterisation and identification of E. coli. The results showed that identification of E. coli strains by each individual method was limited mainly by the extension and quality of reference databases. On the contrary, the polyphasic approach, whereby heterogeneous taxonomic data were combined and weighted, improved the identification results, gave more consistency to the final clustering and provided additional information on the taxonomic structure and phenotypic behaviour of strains, as shown by the close clustering of strains with similar stress resistance patterns.

Use of Potential Probiotic Lactic Acid Bacteria (LAB) Biofilms for the Control of Listeria monocytogenes, Salmonella Typhimurium, and Escherichia coli O157:H7 Biofilms Formation.

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Gómez, Natacha C; Ramiro, Juan M P; Quecan, Beatriz X V; de Melo Franco, Bernadette D G

2016-01-01

Use of probiotic biofilms can be an alternative approach for reducing the formation of pathogenic biofilms in food industries. The aims of this study were (i) to evaluate the probiotic properties of bacteriocinogenic (Lactococcus lactis VB69, L. lactis VB94, Lactobacillus sakei MBSa1, and Lactobacillus curvatus MBSa3) and non-bacteriocinogenic (L. lactis 368, Lactobacillus helveticus 354, Lactobacillus casei 40, and Weissela viridescens 113) lactic acid bacteria (LAB) isolated from Brazilian's foods and (ii) to develop protective biofilms with these strains and test them for exclusion of Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium. LAB were tested for survival in acid and bile salt conditions, surface properties, biosurfactant production, β-galactosidase and gelatinase activity, antibiotic resistance and presence of virulence genes. Most strains survived exposure to pH 2 and 4% bile salts. The highest percentages of auto-aggregation were obtained after 24 h of incubation. Sixty-seven percentage auto-aggregation value was observed in W. viridescens 113 and Lactobacillus curvatus MBSa3 exhibited the highest co-aggregation (69% with Listeria monocytogenes and 74.6% with E. coli O157:H7), while the lowest co-aggregation was exhibited by W. viridescens 113 (53.4% with Listeria monocytogenes and 38% with E. coli O157:H7). Tests for hemolytic activity, bacterial cell adherence with xylene, and drop collapse confirmed the biosurfactant-producing ability of most strains. Only one strain (L. lactis 368) produced β-galactosidase. All strains were negative for virulence genes cob, ccf, cylLL, cylLs, cyllM, cylB, cylA and efaAfs and gelatinase production. The antibiotic susceptibility tests indicated that the MIC for ciprofloxacin, clindamycin, gentamicin, kanamycin, and streptomycin did not exceed the epidemiological cut-off suggested by the European Food Safety Authority. Some strains were resistant to one or more antibiotics and resistance

The isolation, characterization and evaluation of Frankia strains

Energy Technology Data Exchange (ETDEWEB)

Normand, P.; Lalonde, M.; Fortin, J.A.; Chatarpaul, L.

1984-01-01

Osmium tetroxide (OsO/sub 4/) was used as the sterilizing agent to isolate the nitrogen-fixing endophyte Frankia from nodules of host plants. This treatment resulted in a pure culture collection of 250 Frankia isolates from several species of Alnus (crispa, rugosa, viridis, glutinosa, and serrulata) and from other actinorhizal species from Quebec. Frankia isolates from A. viridis and S. canadensis are reported in this document for the first time. Sugars from whole-cell hydrolysates of the organisms were analysed by gas liquid chromatography to identify strains previously screened on the basis of morphology and infectivity. This technique proved especially useful for those isolates whose morphology was atypical (ACN8, ArgN22, SCN9, and SCN10). It was demonstrated that sugar analysis (2-O-methy-D-mannose, in particular) can be important in resolving the taxonomy of Frankia. The nitrogen-fixing efficiency of some Frankia strains was evaluated by inoculating A. crispa. It was found that the efficiency of organisms was not influenced by the host from which they were first isolated. However, significant differences between some isolates of the same provenance occurred. It was also found that those strains of Frankia which do not sporulate were more effective in fixing nitrogen than those which do. It is proposed that the terms type P and type N be used to designate spore positive (Sp/sup +/) and spore negative (Sp/sup -/) Frankia strains, respectively. Further, it is recommended that sporulation type be used as a valid basis for species definition in the genus Frankia. 118 refs., 16 figs., 10 tabs.

Prevalence of Salmonella serovars, Listeria monocytogenes, and Escherichia coli O157:H7 in Mediterranean ready-to-eat meat products in Jordan.

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Osaili, Tareq M; Al-Nabulsi, Anas A; Shaker, Reyad R; Jaradat, Ziad W; Taha, Mohammad; Al-Kherasha, Mohammed; Meherat, Mervet; Holley, Richard

2014-01-01

The presence of Salmonella, Listeria monocytogenes, and Escherichia coli O157:H7 in ready-to-eat (RTE) meat products is considered a major concern for food control authorities worldwide. The aims of this study were to determine (i) the prevalence of Salmonella, L. monocytogenes, and E. coli O157:H7 in Mediterranean RTE chicken and beef (CB) products sold in Jordanian restaurants and (ii) the susceptibility of the isolates to antibiotics. A total of 1,028 samples of various types of RTE CB products (550 RTE chicken and 478 RTE beef products) were analyzed by methods described by the International Organization for Standardization followed by molecular confirmation of the isolates. The VITEK2 automated system was used for testing antibiotic susceptibility of the isolates. The overall prevalence of Salmonella serovars in RTE CB products was 0.5%, with 0.8 and 0.2% in RTE chicken and RTE beef, respectively. The overall prevalence of L. monocytogenes in RTE CB products was 2%, with 2.7 and 1.5% in RTE chicken and RTE beef products, respectively. E. coli O157:H7 was not isolated from any of the tested samples. Multidrug-resistant Salmonella and L. monocytogenes isolates were found. The majority of Salmonella isolates were sensitive to most of the tested antibiotics, and all of the isolates were resistant to more than one antibiotic. Similarly, more than 85% of L. monocytogenes isolates were sensitive to nine antibiotics, and the majority of L. monocytogenes isolates were resistant to fosfomycin and oxacillin.

Efficacy of (+-Lariciresinol to Control Bacterial Growth of Staphylococcus aureus and Escherichia coli O157:H7

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Vivek K. Bajpai

2017-05-01

Full Text Available This study was undertaken to assess the antibacterial potential of a polyphenolic compound (+-lariciresinol isolated from Rubia philippinensis against selected foodborne pathogens Staphylococcus aureus KCTC1621 and Escherichia coli O157:H7. (+-Lariciresinol at the tested concentrations (250 μg/disk evoked a significant antibacterial effect as a diameter of inhibition zones (12.1–14.9 mm with minimum inhibitory concentration (MIC, and minimum bactericidal concentration values of 125–250 and 125–250 μg/mL, respectively. Furthermore, (+-lariciresinol at MIC showed reduction in bacterial cell viabilities, efflux of potassium (K+ ions and release of 260 nm materials against E. coli O157:H7 and S. aureus KCTC1621. Moreover, deteriorated cell wall morphology of E. coli O157:H7 and S. aureus KCTC1621 cells treated with (+-lariciresinol at MIC further confirmed its inhibitory effect against the tested pathogens, suggesting it to be an alternative means of antimicrobials.

Detection of cholera (ctx) and zonula occludens (zot) toxin genes in Vibrio cholerae O1, O139 and non-O1 strains.

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Rivera, I G; Chowdhury, M A; Sanchez, P S; Sato, M I; Huq, A; Colwell, R R; Martins, M T

1995-09-01

Vibrio cholerae O1 and V. cholerae non-O1 strains isolated from environmental samples collected in São Paulo, Brazil, during cholera epidemics and pre-epidemic periods were examined for the presence of toxin genes. V. cholerae O1 strains isolated from clinical samples in Peru and Mexico, and V. cholerae O139 strains from India were also examined for the presence of ctx (cholera toxin gene) and zot (zonula occludens toxin gene) by polymerase chain reaction (PCR). A modified DNA-extraction method applied in this study yielded satisfactory recovery of genomic DNA from vibrios. Results showed that strains of V. cholerae O1 isolated during the preepidemic period were ctx (-)/zot (-) whereas strains isolated during the epidemic were ctx (+)/zot (+). All V. cholerae non-O1 strains tested in the study were ctx (-)/zot (-), whereas all V. cholerae O139 strains were ctx (+)/zot (+). Rapid detection of the virulence genes (ctx and zot) can be achieved by PCR and this can serve as an important tool in the epidemiology and surveillance of V. cholerae.

Validation of the Thermo Scientific SureTect Escherichia coli O157:H7 Real-Time PCR Assay for Raw Beef and Produce Matrixes.

Science.gov (United States)

Cloke, Jonathan; Crowley, Erin; Bird, Patrick; Bastin, Ben; Flannery, Jonathan; Agin, James; Goins, David; Clark, Dorn; Radcliff, Roy; Wickstrand, Nina; Kauppinen, Mikko

2015-01-01

The Thermo Scientific™ SureTect™ Escherichia coli O157:H7 Assay is a new real-time PCR assay which has been validated through the AOAC Research Institute (RI) Performance Tested Methods(SM) program for raw beef and produce matrixes. This validation study specifically validated the assay with 375 g 1:4 and 1:5 ratios of raw ground beef and raw beef trim in comparison to the U.S. Department of Agriculture, Food Safety Inspection Service, Microbiology Laboratory Guidebook (USDS-FSIS/MLG) reference method and 25 g bagged spinach and fresh apple juice at a ratio of 1:10, in comparison to the reference method detailed in the International Organization for Standardization 16654:2001 reference method. For raw beef matrixes, the validation of both 1:4 and 1:5 allows user flexibility with the enrichment protocol, although which of these two ratios chosen by the laboratory should be based on specific test requirements. All matrixes were analyzed by Thermo Fisher Scientific, Microbiology Division, Vantaa, Finland, and Q Laboratories Inc, Cincinnati, Ohio, in the method developer study. Two of the matrixes (raw ground beef at both 1:4 and 1:5 ratios) and bagged spinach were additionally analyzed in the AOAC-RI controlled independent laboratory study, which was conducted by Marshfield Food Safety, Marshfield, Wisconsin. Using probability of detection statistical analysis, no significant difference was demonstrated by the SureTect kit in comparison to the USDA FSIS reference method for raw beef matrixes, or with the ISO reference method for matrixes of bagged spinach and apple juice. Inclusivity and exclusivity testing was conducted with 58 E. coli O157:H7 and 54 non-E. coli O157:H7 isolates, respectively, which demonstrated that the SureTect assay was able to detect all isolates of E. coli O157:H7 analyzed. In addition, all but one of the nontarget isolates were correctly interpreted as negative by the SureTect Software. The single isolate giving a positive result was an E

Rapid detection of E. Coli O157:H7 by IFAST and ATP bioluminescence assay for water analysis

CSIR Research Space (South Africa)

Ngamsom, B

2016-10-01

Full Text Available The present investigation reports isolation and detection of E. coli O157:H7 employing a simple and portable microfluidic device based on immiscible filtration assisted by surface tension (IFAST) and adenosine triphosphate (ATP) bioluminescence...

Longitudinal study of two Irish dairy herds: Low numbers of of Shiga toxin-producing Escherichia coli O157 and O26 super-shedders identified

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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

Detection of viable Escherichia coli O157:H7 in ground beef by propidium monoazide real-time PCR.

Science.gov (United States)

Liu, Yarui; Mustapha, Azlin

2014-01-17

Escherichia coli O157:H7 associated with food has caused many serious public health problems in recent years. However, only viable cells of this pathogen can cause infections, and false-positive detection caused by dead cells can lead to unnecessary product recalls. The objective of this study was to develop and optimize a method that combines propidium monoazide (PMA) staining with real-time PCR to detect only viable cells of E. coli O157:H7 in ground beef. PMA is a DNA intercalating dye that can penetrate compromised membranes of dead cells and bind to cellular DNA, preventing its amplification via a subsequent PCR. Three strains of E. coli O157:H7 (505B, G5310 and C7927) at concentrations of 10(0) to 10(8)CFU/mL were used as live cells. Dead cells were obtained by heating cell suspensions at 85°C for 15 min. Suspensions were treated with PMA and the optimized assay was applied to artificially contaminated ground beef with two different fat contents (10% and 27%). DNA was extracted and amplified by TaqMan® real-time PCR assay targeting the uidA gene for detection of E. coli O157:H7. Plasmid pUC19 was added as an internal amplification control (IAC). A treatment of 25 μM PMA with a 10-min light exposure on ice was sufficient to eliminate DNA from 10(8) dead E. coli O157:H7 cells/mL. The optimized assay could detect as low as 10(2) CFU/mL viable E. coli O157:H7 in pure culture and 10(5) CFU/g in ground beef, in the presence of 10(6)/mL or g of dead cells. With an 8-h enrichment, 1 CFU/g viable E. coli O157:H7 in ground beef was detectable without interference from 10(6) dead cells/g. In conclusion, the PMA real-time PCR could effectively detect viable E. coli O157:H7 without being compromised by dead cells. Copyright © 2013 Elsevier B.V. All rights reserved.

The seaweed fly (Coelopidae) can facilitate environmental survival and transmission of E. coli O157 at sandy beaches.

Science.gov (United States)

Swinscoe, Isobel; Oliver, David M; Gilburn, Andre S; Quilliam, Richard S

2018-06-19

The sustainable management of recreational beaches is essential for minimising risk of human exposure to microbial pathogens whilst simultaneously maintaining valuable ecosystem services. Decaying seaweed on public beaches is gaining recognition as a substrate for microbial contamination, and is a potentially significant reservoir for human pathogens in close proximity to beach users. Closely associated with beds of decaying seaweed are dense populations of the seaweed fly (Coelopidae), which could influence the spatio-temporal fate of seaweed-associated human pathogens within beach environments. Replicated mesocosms containing seaweed inoculated with a bioluminescent strain of the zoonotic pathogen E. coli O157:H7, were used to determine the effects of two seaweed flies, Coelopa frigida and C. pilipes, on E. coli O157:H7 survival dynamics. Multiple generations of seaweed flies and their larvae significantly enhanced persistence of E. coli O157:H7 in simulated wrack habitats, demonstrating that both female and male C. frigida flies are capable of transferring E. coli O157:H7 between individual wrack beds and into the sand. Adult fly faeces can contain significant concentrations of E. coli O157:H7, which suggests they are capable of acting as biological vectors and bridge hosts between wrack habitats and other seaweed fly populations, and facilitate the persistence and dispersal of E. coli O157:H7 in sandy beach environments. This study provides the first evidence that seaweed fly populations inhabiting natural wrack beds contaminated with the human pathogen E. coli O157:H7 have the capacity to amplify the hazard source, and therefore potential transmission risk, to beach users exposed to seaweed and sand in the intertidal zone. The risk to public health from seaweed flies and decaying wrack beds is usually limited by human avoidance behaviour; however, seaweed fly migration and nuisance inland plagues in urban areas could increase human exposure routes beyond the

Determining Thermal Inactivation of Escherichia coli O157:H7 in Fresh Compost by Simulating Early Phases of the Composting Process ▿

OpenAIRE

Singh, Randhir; Kim, Jinkyung; Shepherd, Marion W.; Luo, Feng; Jiang, Xiuping

2011-01-01

A three-strain mixture of Escherichia coli O157:H7 was inoculated into fresh dairy compost (ca. 107 CFU/g) with 40 or 50% moisture and was placed in an environmental chamber (ca. 70% humidity) that was programmed to ramp from room temperature to selected composting temperatures in 2 and 5 days to simulate the early composting phase. The surviving E. coli O157:H7 population was analyzed by direct plating and enrichment. Optimal and suboptimal compost mixes, with carbon/nitrogen (C/N) ratios of...

Identification of E. coli O157:H7 by Using Specific Primers for rfbE and stx2b Genes

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Mostafa Bakhshi

2017-07-01

Sorbitol-MacConkey agar was used to verification of growth ability of selected colonies during PCR. Results: By appearance of the bonds belong to rfbE and stx2B genes on agarose gel, the ability of designed primers in gene detection in samples of E .coli O157:H7 was verified. Colonies which selected during PCR have growth potency on sorbitol-MacConkey agar medium. Conclusion: It was revealed that we can prepare a fast, precise and relative comfortable method for detection of E. coli O157:H7 strain by using PCR technique and specific primers than other available methods.

the occurrence of escherichia coli o157:h7 in market and abattoir

African Journals Online (AJOL)

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Escherichia coli O157:H7 is a newly emerging pathogen frequently associated with the consumption of foods of ... KEY WORDS: E. coli O157:H7, Pathogen, Abattoir, Market, and Infections ..... pathogen. Escherichia coli O157:H7 as a model of.

Search for Enterohaemorrhagic Escherichia coli O157:H7 and ...

African Journals Online (AJOL)

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 and Salmonella enterica are important zoonotic bacteria responsible for enteric infections in humans. The present study investigated the possible role of kittens in the zoonotic transmission of antimicrobial resistant EHEC O157 and Salmonella enterica to human using ...

Survival of bioluminescent Listeria monocytogenes and Escherichia coli O157:H7 in soft cheeses.

Science.gov (United States)

Ramsaran, H; Chen, J; Brunke, B; Hill, A; Griffiths, M W

1998-07-01

Pasteurized and raw milks that had been inoculated at 10(4) cfu/ml with bioluminescent strains of Listeria monocytogenes and Escherichia coli O157:H7 were used in the manufacture of Camembert and Feta cheeses with or without nisin-producing starter culture. Survival of both organisms was determined during the manufacture and storage of Camembert and Feta cheeses at 2 +/- 1 degree C for 65 and 75 d, respectively. Bacterial bioluminescence was used as an indicator to enumerate the colonies plated on selective Listeria agar and on MacConkey agar. Escherichia coli O157:H7 survived the manufacturing process of both cheeses and was present at the end of the storage period in greater numbers than in the initial inoculum. At the end of 75 d of storage, E. coli O157:H7 was found in the brine of Feta cheese. The counts of L. monocytogenes increased as the pH of the Camembert cheese increased, and there were significant differences between the counts from samples taken from the inside and the counts from samples obtained near the surface of the cheese. The Feta cheese that contained nisin was the only cheese in which L. monocytogenes was at the level of the initial inoculum after 75 d of storage.

Quantification of contamination of lettuce by GFP-expressing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium

NARCIS (Netherlands)

Franz, Eelco; Visser, Anna A; Van Diepeningen, Anne D; Klerks, Michel M; Termorshuizen, Aad J; van Bruggen, Ariena H C

The primary objective of this study was to determine the possibility of internalization of GFP-expressing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium (S. Typhimurium) strains MAE 110 (multi-cellular morphology) and 119 (wild type morphology) into lettuce seedlings (Lactuca

Effectiveness of Active Packaging on Control of Escherichia Coli O157:H7 and Total Aerobic Bacteria on Iceberg Lettuce.

Science.gov (United States)

Lu, Haixia; Zhu, Junli; Li, Jianrong; Chen, Jinru

2015-06-01

Contaminated leafy green vegetables have been linked to several outbreaks of human gastrointestinal infections. Antimicrobial interventions that are adoptable by the fresh produce industry for control of pathogen contamination are in great demand. This study was undertaken to evaluate the efficacy of sustained active packaging on control of Escherichia coli O157:H7 and total aerobic bacteria on lettuce. Commercial Iceberg lettuce was inoculated with a 3-strain mixture of E. coli O157:H7 at 10(2) or 10(4) CFU/g. The contaminated lettuce and un-inoculated controls were placed respectively in 5 different active packaging structures. Traditional, nonactive packaging structure was included as controls. Packaged lettuce was stored at 4, 10, or 22 °C for 3 wk and sampled weekly for the population of E. coli O157:H7 and total aerobic bacteria. Results showed that packaging structures with ClO2 generator, CO2 generator, or one of the O2 scavengers effectively controlled the growth of E. coli O157:H7 and total aerobic bacteria under all storage conditions. Packaging structure with the ClO2 generator was most effective and no E. coli O157:H7 was detected in samples packaged in this structure except for those that were inoculated with 4 log CFU/g of E. coli O157:H7 and stored at 22 °C. Packaging structures with an oxygen scavenger and the allyl isothiocyanate generator were mostly ineffective in control of the growth of the bacteria on Iceberg lettuce. The research suggests that some of the packaging structures evaluated in the study can be used to control the presence of foodborne pathogens on leafy green vegetables. © 2015 Institute of Food Technologists®

Antibiotic susceptibility-resistance profiles of super-shed Escherichia coli O157:H7

Science.gov (United States)

Introduction: Escherichia coli O157:H7 (O157) can cause diarrhea and serious secondary sequelae including kidney failure and death in humans. With antibiotics like fosfomycin, colistin and azithromycin, that do not stimulate toxin expression by O157, being considered for treatment of early gastroint...

PCR and ELISA (VIDAS ECO O157® Escherichia coli O157:H7 identification in Minas Frescal cheese commercialized in Goiânia, GO

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Rosangela Nunes Carvalho

2014-01-01

Full Text Available Escherichia coli O157:H7 has been incriminated in food poisoning outbreaks and sporadic cases of hemorrhagic colitis and hemolytic uremic syndrome in many countries. Considering the high susceptibility of Minas Frescal cheese to contamination by E. coli O157:H7, the aim of this study was to determine the occurrence of this pathogen through PCR (Polymerase Chain Reaction and ELISA (VIDAS ECO O157®, bioMérieux, Lyon, France test. Thirty cheese samples manufactured by artisan farmhouse producers were collected from open-air markets in Goiânia and thirty from industries under Federal Inspection located in Goiás State which trade their products in supermarkets in Goiânia. E. coli O157:H7 was detected in 6.67% samples collected in open air markets using ELISA, and 23,33% with PCR. The pathogen was not detected in samples from industries under Federal Inspection.

PCR and ELISA (VIDAS ECO O157®) Escherichia coli O157:H7 identification in Minas Frescal cheese commercialized in Goiânia, GO

Science.gov (United States)

Carvalho, Rosangela Nunes; de Oliveira, Antonio Nonato; de Mesquita, Albenones José; Minafra e Rezende, Cíntia Silva; de Mesquita, Adriano Queiroz; Romero, Rolando Alfredo Mazzoni

2014-01-01

Escherichia coli O157:H7 has been incriminated in food poisoning outbreaks and sporadic cases of hemorrhagic colitis and hemolytic uremic syndrome in many countries. Considering the high susceptibility of Minas Frescal cheese to contamination by E. coli O157:H7, the aim of this study was to determine the occurrence of this pathogen through PCR (Polymerase Chain Reaction) and ELISA (VIDAS ECO O157®, bioMérieux, Lyon, France) test. Thirty cheese samples manufactured by artisan farmhouse producers were collected from open-air markets in Goiânia and thirty from industries under Federal Inspection located in Goiás State which trade their products in supermarkets in Goiânia. E. coli O157:H7 was detected in 6.67% samples collected in open air markets using ELISA, and 23,33% with PCR. The pathogen was not detected in samples from industries under Federal Inspection. PMID:24948907

Inactivation of E. coli O157:H7 on blueberries by electrolyzed water, ultraviolet light, and ozone.

Science.gov (United States)

Kim, Chyer; Hung, Yen-Con

2012-04-01

Increased interest in blueberries due to their nutritional and health benefits has led to an increase in consumption. However, blueberries are consumed mostly raw or minimally processed and are susceptible to microbial contamination like other type of fresh produce. This study was, therefore, undertaken to evaluate the efficacy of electrostatic spray of electrolyzed oxidizing (EO) water, UV light, ozone, and a combination of ozone and UV light in killing Escherichia coli O157:H7 on blueberries. A 5-strain mixture of E. coli O157:H7 were inoculated on the calyx and skin of blueberries and then subjected to the treatments. Electrostatic EO water spray reduced initial populations of E. coli O157:H7 by only 0.13 to 0.24 log CFU/g and 0.88 to 1.10 log CFU/g on calyx and skin of blueberries, respectively. Ozone treatment with 4000 mg/L reduced E. coli O157:H7 by only 0.66 and 0.72 log CFU/g on calyx and skin of blueberries, respectively. UV light at 20 mW/cm² for 10 min was the most promising single technology and achieved 2.14 and greater than 4.05 log reductions of E. coli O157:H7 on the calyx and skin of blueberries, respectively. The combination treatment of 1 min ozone and followed by a 2 min UV achieved more than 1 and 2 log additional reductions on blueberry calyx than UV or ozone alone, respectively. Outbreaks of foodborne illnesses have been associated with consumption of fresh produce. Many methods for removing pathogens as well as minimizing their effect on quality of treated produce have been investigated. UV technology and its combination with ozone used in this study to inactive E. coli O157:H7 on blueberries was found effective. Results from this study may help producers and processors in developing hurdle technologies for the delivery of safer blueberries to consumers. © 2012 Institute of Food Technologists®

RAPD-PCR typing of Yersinia enterocolitica (Enterobacteriaceae O:3 serotype strains isolated from pigs and humans

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Tereza Cristina A. Leal

1999-09-01

Full Text Available Sixteen strains of Yersinia enterocolitica serotype O:3, isolated from apparently healthy pigs collected in Rio de Janeiro, and four human strains of serotypes O:4, O:5, O:6 and O:13 were analyzed by RAPD-PCR. The strains were grouped into five genotypic profiles according to the amplification patterns obtained with three random primers. Fifteen of the 16 pig strains had identical amplification patterns, which was named genotypic profile 1. The one different profile was named genotypic profile 2. Genotypic profile 1 was also exhibited by the O:6 human serotype strain. The O:4 and O:13 human serotype strains showed similar amplification profiles with two primers. However, the third primer induced a distinct profile in each strain. Therefore, these two strains were placed into genotypic profile 3 and 4, respectively. Each primer produced a completely different amplification profile in the O:5 human serotype strain; therefore, it was named genotypic profile 5. The presence or absence of plasmids in the strains studied did not affect the amplification results. These results show that genetic variations can exist within a serotype, and strains of different serotypes can exhibit the same amplification profile when compared using other primers.Foram utilizados três "primers" aleatórios para caracterizar pela técnica RAPD-PCR 16 cepas de Yersinia enterocolitica do sorotipo O:3, isoladas de suínos sadios do Rio de Janeiro. Pelos resultados dos padrões de amplificação, as 16 cepas dos suínos e as 4 cepas humanas usadas como referência (sorotipos O:4, O:5, O:6 e O:13 foram agrupadas em 5 perfis genotípicos. Quinze cepas de suínos apresentaram um padrão de amplificação idêntico (perfil genotípico 1 e somente uma apresentou um perfil de amplificação diferente (perfil genotípico 2. O mesmo padrão de amplificação do perfil genotípico 1 foi também observado em uma cepa humana do sorotipo O:6. As cepas humanas dos sorotipos O:4 e O:13

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.

VTEC O157 subtypes associated with the most severe clinical symptoms in humans constitute a minor part of VTEC 0157 isolates from Danish Cattle

DEFF Research Database (Denmark)

Roldgaard, Bemt Bjørn; Scheutz, Flemming; Boel, Jeppe

2004-01-01

-positive VTEC 0 157 isolates (63 of bovine origin and 86 from human clinical cases) isolated between 1987 and 2001. All were analysed by vtx-PCR-RFLP and phage typing. The vtx-PCR-RFLP showed that isolates carrying the vtx2 gene was more than four times as prevalent among the human clinical isolates (55...

Effects of Plant-Derived Extracts, Other Antimicrobials, and Their Combinations against Escherichia coli O157:H7 in Beef Systems.

Science.gov (United States)

Ko, Kyung Yuk; Geornaras, Ifigenia; Paik, Hyun-Dong; Kim, Kee-Tae; Sofos, John N

2015-06-01

The antimicrobial effects of thyme oil (TO), grapefruit seed extract (GSE), and basil essential oil, alone or in combination with cetylpyridinium chloride (CPC), sodium diacetate, or lactic acid, were evaluated against Escherichia coli O157:H7 in a moisture-enhanced beef model system. The model system was composed of a nonsterile beef homogenate to which NaCl (0.5%) and sodium tripolyphosphate (0.25%) were added, together with the tested antimicrobial ingredients. Beef homogenate treatments were inoculated (ca. 3 log CFU/ml) with rifampin-resistant E. coli O157:H7 (eight-strain mixture) and incubated at 15 °C (48 h). The most effective individual treatments were TO (0.25 or 0.5%) and GSE (0.5 or 1.0%), which immediately reduced (P extracts with CPC (0.02 or 0.04%) and sodium diacetate (0.25%) had an additive effect with respect to antibacterial activity. In a second study, antimicrobial interventions were evaluated for their efficacy in reducing surface contamination of E. coli O157:H7 on beef cuts and to determine the effect of these surface treatments on subsequent internalization of the pathogen during blade tenderization. Beef cuts (10 by 8 by 3.5 cm) were inoculated (ca. 4 log CFU/g) on one side with the rifampin-resistant E. coli O157:H7 strain mixture and were then spray treated (20 lb/in(2), 10 s) with water, GSE (5 and 10%), lactic acid (5%), or CPC (5%). Untreated (control) and spray-treated surfaces were then subjected to double-pass blade tenderization. Surface contamination (4.4 log CFU/g) of E. coli O157:H7 was reduced (P < 0.05) to 3.4 (5% CPC) to 4.1 (water or 5% GSE) log CFU/g following spray treatment. The highest and lowest transfer rates of pathogen cells from the surface to deeper tissues of blade-tenderized sections were obtained in the untreated control and CPC-treated samples, respectively.

Characterization of high exopolysaccharide-producing Lactobacillus strains isolated from mustard pickles for potential probiotic applications.

Science.gov (United States)

Huang, Jing-Yao; Kao, Cheng-Yen; Liu, We-Sin; Fang, Tony J

2017-06-01

The aim of this study was to characterize high exopolysaccharide (EPS)-producing lactic acid bacteria (LAB) isolated from mustard pickles in Taiwan for potential probiotic applications. Among 39 collected LAB strains, four most productive EPS-producing strains were selected for further analysis. Comparative analyses of 16S rDNA genes rpoA and pheS sequences demonstrated that these strains were members of Lactobacillus plantarum-group (LPG). NCD 2, NLD 4, SLC 13, and NLD 16 showed survival rates of 95.83% ± 0.49%, 95.07% ± 0.64%, 105.84% ± 0.82%, and 99.65% ± 0.31% under simulated gastrointestinal conditions, respectively. No cytotoxic effects on macrophage RAW 264.7 cells were observed when they were treated with a low dose (1 μg/ml) of stimulants extracted from the tested LAB strains. The production of nitric oxide in RAW 264.7 cells incubated with various LAB stimulants showed a dose-dependent increase. Among the four strains, SLC 13 showed higher inhibitory activity on growth of Enterococcus faecalis (BCRC 12302) and Yersinia enterocolitica (BCRC 10807). NLD 4 showed strong inhibitory activity against Escherichia coli O157:H7 (ATCC 43894) as compared with the other three strains. In summary, our results suggest that Lactobacillus pentosus SLC 13 may be a good candidate for probiotic applications and for development of antibacterial compounds. [Int Microbiol 20(2):75-84 (2017)]. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

Transcriptome Changes of Escherichia coli, Enterococcus faecalis, and Escherichia coli O157:H7 Laboratory Strains in Response to Photo-Degraded DOM

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Adelumola Oladeinde

2018-05-01

Full Text Available In this study, we investigated gene expression changes in three bacterial strains (Escherichia coli C3000, Escherichia coli O157:H7 B6914, and Enterococcus faecalis ATCC 29212, commonly used as indicators of water quality and as control strains in clinical, food, and water microbiology laboratories. Bacterial transcriptome responses from pure cultures were monitored in microcosms containing water amended with manure-derived dissolved organic matter (DOM, previously exposed to simulated sunlight for 12 h. We used RNA sequencing (RNA-seq and quantitative real-time reverse transcriptase (qRT-PCR to compare differentially expressed temporal transcripts between bacteria incubated in microcosms containing sunlight irradiated and non-irradiated DOM, for up to 24 h. In addition, we used whole genome sequencing simultaneously with RNA-seq to identify single nucleotide variants (SNV acquired in bacterial populations during incubation. These results indicate that E. coli and E. faecalis have different mechanisms for removal of reactive oxygen species (ROS produced from irradiated DOM. They are also able to produce micromolar concentrations of H2O2 from non-irradiated DOM, that should be detrimental to other bacteria present in the environment. Notably, this study provides an assessment of the role of two conjugative plasmids carried by the E. faecalis and highlights the differences in the overall survival dynamics of environmentally-relevant bacteria in the presence of naturally-produced ROS.

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.

O-serotyping of Escherichia coli Strains isolated from Patients With Urinary Tract Infection in Southeast of Iran

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Ahmad Rashki

2014-11-01

Full Text Available Background: Uropathogenic Escherichia coli (UPEC O-serogroups with their phylogenetic background are the most prevalent causes of urinary tract infections (UTIs. Objectives: The association of O types with phylogenetic background was assessed among E. coli isolates collected from patients with UTI. Patients and Methods: In this study, 186 patients with UTI, referred to two hospitals affiliated to Zabol University of Medical Sciences in southeast of Iran, were enrolled during January to July 2013. Phylogenetic groups and serotyping were performed using multiplex-PCR method. Results: A total of 100 E. coli strains were isolated from the urine samples. The most common types of O antigens were O2 (16.43%, O6 (16.43% and O18 (13.69%. The phylogenetic analysis showed that 63 O-antigen-positive isolates were mainly segregated from the phylogenetic group B2 (56% and the substantial prevalence (30% belonged to the phylogenetic group D. Conclusions: This was the first report of E. coli serotyping in patient with UTI from southeast of Iran as well as investigation of their relation with phylogenetic pattern by multiplex-PCR. Further studies from other parts of Iran and on other serotypes are recommended.

Saltelli Global Sensitivity Analysis and Simulation Modelling to Identify Intervention Strategies to Reduce the Prevalence of Escherichia coli O157 Contaminated Beef Carcasses.

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Victoria J Brookes

Full Text Available Strains of Shiga-toxin producing Escherichia coli O157 (STEC O157 are important foodborne pathogens in humans, and outbreaks of illness have been associated with consumption of undercooked beef. Here, we determine the most effective intervention strategies to reduce the prevalence of STEC O157 contaminated beef carcasses using a modelling approach.A computational model simulated events and processes in the beef harvest chain. Information from empirical studies was used to parameterise the model. Variance-based global sensitivity analysis (GSA using the Saltelli method identified variables with the greatest influence on the prevalence of STEC O157 contaminated carcasses. Following a baseline scenario (no interventions, a series of simulations systematically introduced and tested interventions based on influential variables identified by repeated Saltelli GSA, to determine the most effective intervention strategy.Transfer of STEC O157 from hide or gastro-intestinal tract to carcass (improved abattoir hygiene had the greatest influence on the prevalence of contaminated carcases. Due to interactions between inputs (identified by Saltelli GSA, combinations of interventions based on improved abattoir hygiene achieved a greater reduction in maximum prevalence than would be expected from an additive effect of single interventions. The most effective combination was improved abattoir hygiene with vaccination, which achieved a greater than ten-fold decrease in maximum prevalence compared to the baseline scenario.Study results suggest that effective interventions to reduce the prevalence of STEC O157 contaminated carcasses should initially be based on improved abattoir hygiene. However, the effect of improved abattoir hygiene on the distribution of STEC O157 concentration on carcasses is an important information gap-further empirical research is required to determine whether reduced prevalence of contaminated carcasses is likely to result in reduced

Influence of surface polysaccharides of Escherichia coli O157:H7 on plant defense response and survival of the human enteric pathogen on Arabidopsis thaliana and lettuce (Lactuca sativa).

Science.gov (United States)

Jang, Hyein; Matthews, Karl R

2018-04-01

This study aimed to determine the influence of bacterial surface polysaccharides (cellulose, colanic acid, and lipopolysaccharide; LPS) on the colonization or survival of Escherichia coli O157:H7 on plants and the plant defense response. Survival of E. coli O157:H7 were evaluated on Arabidopsis thaliana and romaine lettuce as a model plant and an edible crop (leafy vegetable), respectively. The population of the wild-type strain of E. coli O157:H7 on Arabidopsis plants and lettuce was significantly (P lettuce regardless of day post-inoculation. Copyright © 2017 Elsevier Ltd. All rights reserved.

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.

Disruption of rcsB by a duplicated sequence in a curli-producing Escherichia coli O157:H7 results in differential gene expression in relation to biofilm formation, stress responses, and metabolism

Science.gov (United States)

Background: Escherichia coli O157:H7 (O157) strain 86-24, linked to a 1986 disease outbreak, displays biofilm- and curli-negative phenotypes that are correlated with the lack of Congo red (CR) binding and formation of white colonies (CR negative) on a CR negative containing medium. However, on a CR ...

Antibacterial activity of selected plant essential oils against Escherichia coli O157:H7.

Science.gov (United States)

Burt, S A; Reinders, R D

2003-01-01

To quantify the antibacterial properties of five essential oils (EO) on a non-toxigenic strain of Escherichia coli O157:H7 in the presence and absence of a stabilizer and an emulsifier and at three different temperatures. Five EOs known to exhibit antibacterial properties were screened by disc diffusion assay and the most active were selected for further study in microdilution colorimetric assays. Oregano (Origanum vulgare) and thyme (Thymus vulgaris; light and red varieties) EO had the strongest bacteriostatic and bactericidal properties, followed by bay (Pimenta racemosa) and clove bud (Eugenia caryophyllata synonym: Syzygium aromaticum) EO. Oregano oil was colicidal at 625 microl l(-1) at 10, 20 and 37 degrees C. The addition of 0.05% (w/v) agar as stabilizer reinforced the antibacterial properties, particularly at 10 degrees C, whereas 0.25% (w/v) lecithin reduced antibacterial activity. Scanning electron micrographs showed extensive morphological changes to treated cells. Oregano and thyme EO possess significant in vitro colicidal and colistatic properties, which are exhibited in a broad temperature range and substantially improved by the addition of agar as stabilizer. Bay and clove bud EO are less active. Lecithin diminished antibacterial properties. The bactericidal concentration of oregano EO irreversibly damaged E. coli O157:H7 cells within 1 min. Oregano and light thyme EO, particularly when enhanced by agar stabilizer, may be effective in reducing the number or preventing the growth of E. coli O157:H7 in foods.

Quantitative transfer of Escherichia coli O157:H7 to equipment during small-scale production of fresh-cut leafy greens.

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Buchholz, Annemarie L; Davidson, Gordon R; Marks, Bradley P; Todd, Ewen C D; Ryser, Elliot T

2012-07-01

Postharvest contamination and subsequent spread of Escherichia coli O157:H7 can occur during shredding, conveying, fluming, and dewatering of fresh-cut leafy greens. This study quantified E. coli O157:H7 transfer from leafy greens to equipment surfaces during simulated small-scale commercial processing. Three to five batches (22.7 kg) of baby spinach, iceberg lettuce, and romaine lettuce were dip inoculated with a four-strain cocktail of avirulent, green fluorescent protein-labeled, ampicillinresistant E. coli O157:H7 to contain ∼10(6), 10(4), and 10(2) CFU/g, and then were processed after 1 h of draining at ∼23°C or 24 h of storage at 4°C. Lettuce was shredded using an Urschel TransSlicer at two different blade and belt speeds to obtain normal (5 by 5 cm) and more finely shredded (0.5 by 5 cm) lettuce. Thereafter, the lettuce was step conveyed to a flume tank and was washed and then dried using a shaker table and centrifugal dryer. Product (25-g) and water (40-ml) samples were collected at various points during processing. After processing, product contact surfaces (100 cm(2)) on the shredder (n = 14), conveyer (n = 8), flume tank (n = 11), shaker table (n = 9), and centrifugal dryer (n = 8) were sampled using one-ply composite tissues. Sample homogenates diluted in phosphate or neutralizing buffer were plated, with or without prior 0.45- m m membrane filtration, on Trypticase soy agar containing 0.6% yeast extract supplemented with 100 ppm of ampicillin to quantify green fluorescent protein-labeled E. coli O157:H7 under UV light. During leafy green processing, ∼90% of the E. coli O157:H7 inoculum transferred to the wash water. After processing, E. coli O157:H7 populations were highest on the conveyor and shredder (Ptransfer.

Experimental mouse lethality of Escherichia coli strains isolated from free ranging Tibetan yaks.

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Rehman, Mujeeb Ur; Zhang, Hui; Wang, Yajing; Mehmood, Khalid; Huang, Shucheng; Iqbal, Muhammad Kashif; Li, Jiakui

2017-08-01

The present study has examined the virulence potential of Escherichia coli isolates harboring at least one virulence gene (associated with ExPEC or InPEC pathotype and belonging to different phylogenetic groups: A, B1, B2 or D), isolated from free ranging Tibetan yak feces. The E. coli isolates (n = 87) were characterized for different serogroups and a mouse model of subcutaneous-infection was used to envisage the virulence within these E. coli strains. Of the 87 E. coli isolates examined, 23% of the E. coli isolates caused lethal infections in a mouse model of subcutaneous infection and were classified as killer. Moreover, the majority of the killer strains belonged to phylogroup A (65%) and serogroup O 60 or O 101 (35%). Phylogroup B1, serogroups O 60 and O 101 were statistically associated with the killer status (P 1) were observed between the killer status isolates and all other bacterial virulence traits. This study comprises the first report on the virulence potential of E. coli strains isolated from free-ranging Tibetan yaks feces. Our findings suggest that pathogenic E. coli of free ranging yaks is highly worrisome, as these feces are used as manures by farmers and therewith pose a health risk to humans upon exposure. Copyright © 2017 Elsevier Ltd. All rights reserved.

Isolation of pathogenic Yersinia enterocolitica strains from different sources in Izmir region, Turkey.

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Bozcal, Elif; Uzel, Atac; Aydemir, Sohret; Skurnik, Mikael

2015-11-01

Yersinia enterocolitica is a foodborne pathogen that is very rarely encountered in Turkey. In this work, several human, porcine, and environmental samples collected from Izmir region in Turkey were examined for the presence of Y. enterocolitica using different cultivation and enrichment methods. A total of nine pathogenic Y. enterocolitica strains were isolated; five strains from pig stool and manure samples and four strains from waste water samples. On the other hand, no Y. enterocolitica was isolated from human diarrheal stool samples (n = 102) and from 12 gulf, canal, municipal pool, and well water samples. Biochemical and serological characterization of the nine Y. enterocolitica strains revealed that they belonged to three different bioserotypes: 4/O:3, 2/O:9, and 2/O:5,27. All the strains were deemed pathogenic based on virulence factor-specific PCR analysis. Detection of pathogenic Y. enterocolitica strains from the pig and waste water samples from the Izmir region indicates that Y. enterocolitica is a potential risk for public health.

Growth media simulating ileal and colonic environments affect the intracellular proteome and carbon fluxes of enterohemorrhagic Escherichia coli O157:H7 strain EDL933.

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Polzin, Sabrina; Huber, Claudia; Eylert, Eva; Elsenhans, Ines; Eisenreich, Wolfgang; Schmidt, Herbert

2013-06-01

In this study, the intracellular proteome of Escherichia coli O157:H7 strain EDL933 was analyzed by two-dimensional gel electrophoresis and matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) spectrometry after growth in simulated ileal environment media (SIEM) and simulated colonic environment media (SCEM) under aerobic and microaerobic conditions. Differentially expressed intracellular proteins were identified and allocated to functional protein groups. Moreover, metabolic fluxes were analyzed by isotopologue profiling with [U-(13)C(6)]glucose as a tracer. The results of this study show that EDL933 responds with differential expression of a complex network of proteins and metabolic pathways, reflecting the high metabolic adaptability of the strain. Growth in SIEM and SCEM is obviously facilitated by the upregulation of nucleotide biosynthesis pathway proteins and could be impaired by exposition to 50 µM 6-mercaptopurine under aerobic conditions. Notably, various stress and virulence factors, including Shiga toxin, were expressed without having contact with a human host.

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

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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

[Virulence markers of Escherichia coli O1 strains].

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Makarova, M A; Kaftyreva, L A; Grigor'eva, N S; Kicha, E V; Lipatova, L A

2011-01-01

To detect virulence genes in clinical isolates of Escherichia coli O1 using polymerase chain reaction (PCR). One hundred and twenty strains of E.coli O1 strains isolated from faeces of patients with acute diarrhea (n = 45) and healthy persons (n = 75) were studied. PCR with primers for rfb and fliC genes, which control synthesis of O- and H- antigens respectively, was used. Fourteen virulence genes (pap, aaf, sfa, afa, eaeA, bfpA, ial, hly, cnf, stx1, stx2, lt, st, and aer) were detected by PCR primers. K1-antigen was determined by Pastorex Meningo B/E. coli O1 kit (Bio-Rad). rfb gene controlling O-antigen synthesis in serogroup O1 as well as fliC gene controlling synthesis of H7 and K1 antigens were detected in all strains. Thus all E. coli strains had antigenic structure O1:K1 :H-:F7. Virulence genes aafl, sfa, afa, eaeA, bfpA, ial, hly, cnf, stx1, stx2, lt, and st were not detected. All strains owned pap and aer genes regardless of the presence of acute diarrhea symptoms. It was shown that E. coli O1:KI:H-:F7 strains do not have virulence genes which are characteristic for diarrhea-causing Escherichia. In accordance with the presence of pap and aer genes they could be attributed to uropathogenic Escherichia (UPEC) or avian-pathogenic Escherichia (APEC). It is necessary to detect virulence factors in order to determine E. coli as a cause of intestinal infection.

Recto-anal junction (RAJ) microbiota composition in Escherichia coli O157:H7 shedding cattle

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Introduction: Cattle are the asymptomatic reservoirs of Escherichia coli O157:H7 (O157) that tend to preferentially colonize the bovine recto-anal junction (RAJ). Therefore, understanding the taxonomic profile, microbial diversity, and microbiota-O157 interactions at the RAJ could give insights into...

Molecular analyses of Vibrio cholerae O1 clinical strains, including new nontoxigenic variants isolated in Mexico during the Cholera epidemic years between 1991 and 2000.

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Lizárraga-Partida, Marcial Leonardo; Quilici, Marie-Laure

2009-05-01

We studied the evolution of Vibrio cholerae O1 during the 1991 to 2000 cholera epidemic in Mexico by biochemical, serological, and molecular characterization of strains collected during this period. Strains were divided into toxigenic and nontoxigenic groups according to the presence or absence of genes encoding cholera toxin. As previously reported, we characterized two populations among toxigenic strains, which were present from the first year of the epidemic. BglI rRNA analysis revealed that these strains had ribotype profiles, denoted M5 and M6 in our study, that were identical to those previously designated Koblavi B5 or Popovic 5 and Popovic 6a or Tamayo B21a, respectively. Ribotype M5 was isolated between 1991 and 1993. This ribotype had a low level of genetic variation as detected by pulsed-field gel electrophoresis (PFGE). Ribotype M6 persisted from 1991 to 2000. However, PFGE profiles suggested that two epidemiologically unrelated strains coexisted within this single ribotype from 1995 until the end of the epidemic. We identified three new BglI ribotypes, Mx1, Mx2, and Mx3, from nontoxigenic V. cholerae O1 strains isolated between 1998 and 2000; one of them grouped strains positive for the toxin-coregulated pilus island. They differed from nontoxigenic clones isolated in Latin America and on the U.S. Gulf Coast and are probably autochthonous Mexican V. cholerae O1 variants. Most of these new variants were isolated from states surrounding the Gulf of Mexico, where the highest incidence of cholera in the country was recorded. Thus, the Mexican Gulf Coast, like the U.S. Gulf Coast, may act as an environmental reservoir of V. cholerae O1.

Microarray-Based Screening of Differentially Expressed Genes of E. coli O157:H7 Sakai during Preharvest Survival on Butterhead Lettuce

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Inge Van der Linden

2016-01-01

Full Text Available Numerous outbreaks of Escherichia coli O157:H7 have been linked to the consumption of leafy vegetables. However, up to the present, little has been known about E. coli O157:H7’s adaptive responses to survival on actively growing (and thus responsive plants. In this study, whole genome transcriptional profiles were generated from E. coli O157:H7 cells (isolate Sakai, stx- one hour and two days after inoculation on the leaves of growing butterhead lettuce, and compared with an inoculum control. A total of 273 genes of E. coli O157:H7 Sakai (5.04% of the whole genome were significantly induced or repressed by at least two-fold (p < 0.01 in at least one of the analyzed time points in comparison with the control. Several E. coli O157:H7 genes associated with oxidative stress and antimicrobial resistance were upregulated, including the iron-sulfur cluster and the multiple antibiotic resistance (mar operon, whereas the Shiga toxin virulence genes were downregulated. Nearly 40% of the genes with significantly different expression were poorly characterized genes or genes with unknown functions. These genes are of special interest for future research as they may play an important role in the pathogens’ adaptation to a lifestyle on plants. In conclusion, these findings suggest that the pathogen actively interacts with the plant environment by adapting its metabolism and responding to oxidative stress.

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

Microbiota response to Escherichia coli O157:H7 colonization in cattle

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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...

High temperature in combination with UV irradiation enhances horizontal transfer of stx2 gene from E. coli O157:H7 to non-pathogenic E. coli.

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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.

Potential application of high hydrostatic pressure to eliminate Escherichia coli O157:H7 on alfalfa sprouted seeds.

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Neetoo, Hudaa; Ye, Mu; Chen, Haiqiang

2008-12-10

Sprouts eaten raw are increasingly being perceived as hazardous foods as they have been implicated in Escherichia coli O157:H7 outbreaks where the seeds were found to be the likely source of contamination. The objective of our study was to evaluate the potential of using high hydrostatic pressure (HHP) technology for alfalfa seed decontamination. Alfalfa seeds inoculated with a cocktail of five strains of E. coli O157:H7 were subjected to pressures of 500 and 600 MPa for 2 min at 20 degrees C in a dry or wet (immersed in water) state. Immersing seeds in water during pressurization considerably enhanced inactivation of E. coli O157:H7 achieving reductions of 3.5 log and 5.7 log at 500 and 600 MPa, respectively. When dry seeds were pressurized, both pressure levels reduced the counts by 5 log reduction in the population was achieved when 600 MPa was applied for durations of > or =6 min although survivors were still detected by enrichment. When the pressure was stepped up to 650 MPa, the threshold time required to achieve complete elimination was 15 min. Un-inoculated seeds pressure-treated at 650 MPa for 15 min at 20 degrees C successfully sprouted achieving a germination rate identical to untreated seeds after eight days of sprouting. These results therefore demonstrate the promising application of HHP on alfalfa seeds to eliminate the risk of E. coli O157:H7 infections associated with consumption of raw alfalfa sprouts.

Characterization of a clinical Vibrio cholerae O139 isolate from Mexico.

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Parveen, Salina; Farrah, Samuel R; Gonzalez-Bonilla, Celia; Zamudio, Altagracia V; Tamplin, Mark L

2003-01-01

Pathogenic strains of Vibrio cholerae O139 possess the cholera toxin A subunit (ctxA) gene as well as the gene for toxin co-regulated pili (tcpA). We report the isolation of a ctxA-negative, tcpA-negative V. cholerae O139 strain (INDREI) from a patient in Mexico diagnosed with gastrointestinal illness. Certain phenotypic characteristics of this strain were identical to those of V. cholerae O1 biotype El Tor. Unlike ctxA-positive V. cholerae O139 strains, this strain was sensitive to a wide panel of antibiotics, including ampicillin, chloramphenicol, ciprofloxacin, gentamicin, furazolidone, nalidixic acid, nitrofurantoin, tetracycline, trimethoprim-sulfamethoxazole, and streptomycin, but was resistant to polymyxin B. Ribotype and pulsed-field gel electrophoresis profiles of INDRE1 differed from those of ctxA-positive V. cholerae O139 and other V. cholerae strains. Phenotypic characteristics of the Mexico strain were similar to those reported for V. cholerae O139 isolates from Argentina and Sri Lanka.

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

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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.

The highly virulent 2006 Norwegian EHEC O103:H25 outbreak strain is related to the 2011 German O104:H4 outbreak strain.

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Trine M L'Abée-Lund

Full Text Available In 2006, a severe foodborne EHEC outbreak occured in Norway. Seventeen cases were recorded and the HUS frequency was 60%. The causative strain, Esherichia coli O103:H25, is considered to be particularly virulent. Sequencing of the outbreak strain revealed resemblance to the 2011 German outbreak strain E. coli O104:H4, both in genome and Shiga toxin 2-encoding (Stx2 phage sequence. The nucleotide identity between the Stx2 phages from the Norwegian and German outbreak strains was 90%. During the 2006 outbreak, stx(2-positive O103:H25 E. coli was isolated from two patients. All the other outbreak associated isolates, including all food isolates, were stx-negative, and carried a different phage replacing the Stx2 phage. This phage was of similar size to the Stx2 phage, but had a distinctive early phage region and no stx gene. The sequence of the early region of this phage was not retrieved from the bacterial host genome, and the origin of the phage is unknown. The contaminated food most likely contained a mixture of E. coli O103:H25 cells with either one of the phages.

The Escherichia coli O157:H7 bovine rumen fluid proteome reflects adaptive bacterial responses

OpenAIRE

Kudva, Indira T; Stanton, Thaddeus B; Lippolis, John D

2014-01-01

Background To obtain insights into Escherichia coli O157:H7 (O157) survival mechanisms in the bovine rumen, we defined the growth characteristics and proteome of O157 cultured in rumen fluid (RF; pH 6.0-7.2 and low volatile fatty acid content) obtained from rumen-fistulated cattle fed low protein content “maintenance diet” under diverse in vitro conditions. Results Bottom-up proteomics (LC-MS/MS) of whole cell-lysates of O157 cultured under anaerobic conditions in filter-sterilized RF (fRF; d...

Comparison of Diversities of Escherichia coli O157 Shed from a Cohort of Spring-Born Beef Calves at Pasture and in Housing

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Vali, Leila; Pearce, Michael C.; Wisely, Karen A.; Hamouda, Ahmed; Knight, Hazel I.; Smith, Alastair W.; Amyes, Sebastian G. B.

2005-01-01

A cohort of spring-born beef calves demonstrated limited genetic and phenotypic diversity of Escherichia coli O157 when kept in a state of isolation. Despite this, there was a difference in the pulsed-field gel electrophoresis and phage types of isolates shed by cattle at pasture compared with those shed by the same cattle when weaned and housed. PMID:15746371

Molecular Analyses of Vibrio cholerae O1 Clinical Strains, Including New Nontoxigenic Variants Isolated in Mexico during the Cholera Epidemic Years between 1991 and 2000▿

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Lizárraga-Partida, Marcial Leonardo; Quilici, Marie-Laure

2009-01-01

We studied the evolution of Vibrio cholerae O1 during the 1991 to 2000 cholera epidemic in Mexico by biochemical, serological, and molecular characterization of strains collected during this period. Strains were divided into toxigenic and nontoxigenic groups according to the presence or absence of genes encoding cholera toxin. As previously reported, we characterized two populations among toxigenic strains, which were present from the first year of the epidemic. BglI rRNA analysis revealed that these strains had ribotype profiles, denoted M5 and M6 in our study, that were identical to those previously designated Koblavi B5 or Popovic 5 and Popovic 6a or Tamayo B21a, respectively. Ribotype M5 was isolated between 1991 and 1993. This ribotype had a low level of genetic variation as detected by pulsed-field gel electrophoresis (PFGE). Ribotype M6 persisted from 1991 to 2000. However, PFGE profiles suggested that two epidemiologically unrelated strains coexisted within this single ribotype from 1995 until the end of the epidemic. We identified three new BglI ribotypes, Mx1, Mx2, and Mx3, from nontoxigenic V. cholerae O1 strains isolated between 1998 and 2000; one of them grouped strains positive for the toxin-coregulated pilus island. They differed from nontoxigenic clones isolated in Latin America and on the U.S. Gulf Coast and are probably autochthonous Mexican V. cholerae O1 variants. Most of these new variants were isolated from states surrounding the Gulf of Mexico, where the highest incidence of cholera in the country was recorded. Thus, the Mexican Gulf Coast, like the U.S. Gulf Coast, may act as an environmental reservoir of V. cholerae O1. PMID:19213700

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.

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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

Comparative genomic analysis shows that avian pathogenic Escherichia coli isolate IMT5155 (O2:K1:H5; ST complex 95, ST140 shares close relationship with ST95 APEC O1:K1 and human ExPEC O18:K1 strains.

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Xiangkai Zhu Ge

Full Text Available Avian pathogenic E. coli and human extraintestinal pathogenic E. coli serotypes O1, O2 and O18 strains isolated from different hosts are generally located in phylogroup B2 and ST complex 95, and they share similar genetic characteristics and pathogenicity, with no or minimal host specificity. They are popular objects for the study of ExPEC genetic characteristics and pathogenesis in recent years. Here, we investigated the evolution and genetic blueprint of APEC pathotype by performing phylogenetic and comparative genome analysis of avian pathogenic E. coli strain IMT5155 (O2:K1:H5; ST complex 95, ST140 with other E. coli pathotypes. Phylogeny analyses indicated that IMT5155 has closest evolutionary relationship with APEC O1, IHE3034, and UTI89. Comparative genomic analysis showed that IMT5155 and APEC O1 shared significant genetic overlap/similarities with human ExPEC dominant O18:K1 strains (IHE3034 and UTI89. Furthermore, the unique PAI I5155 (GI-12 was identified and found to be conserved in APEC O2 serotype isolates. GI-7 and GI-16 encoding two typical T6SSs in IMT5155 might be useful markers for the identification of ExPEC dominant serotypes (O1, O2, and O18 strains. IMT5155 contained a ColV plasmid p1ColV5155, which defined the APEC pathotype. The distribution analysis of 10 sequenced ExPEC pan-genome virulence factors among 47 sequenced E. coli strains provided meaningful information for B2 APEC/ExPEC-specific virulence factors, including several adhesins, invasins, toxins, iron acquisition systems, and so on. The pathogenicity tests of IMT5155 and other APEC O1:K1 and O2:K1 serotypes strains (isolated in China through four animal models showed that they were highly virulent for avian colisepticemia and able to cause septicemia and meningitis in neonatal rats, suggesting zoonotic potential of these APEC O1:K1 and O2:K1 isolates.

Fate of Escherichia coli O157:H7, Salmonella Enteritidis and Listeria monocytogenes during storage of fermented green table olives in brine.

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Argyri, Anthoula A; Lyra, Efstathia; Panagou, Efstathios Z; Tassou, Chrysoula C

2013-10-01

The survival of Escherichia coli O157:H7, Salmonella Enteritidis and Listeria monocytogenes during the storage of fermented green table olives cv. Halkidiki in brine was studied in parallel with the evolution of lactic acid bacteria (LAB), yeasts and pH. The olives were previously fermented with a starter culture (a potential probiotic strain of Lactobacillus pentosus B281--starter process) or with the indigenous microbiota (control). After the end of fermentation, olives were placed in brine, inoculated with a cocktail of 5 strains of E. coli O157:H7, 5 strains of L. monocytogenes and 4 strains of S. Enteritidis, with a final concentration in the brine of ca. 7.0 log CFU/ml, and subsequently packaged in polyethylene pouches and stored at 20 °C. The population of E. coli O157:H7 reduced gradually and was detected in the brine until the 27th day of storage in both cases (i.e., starter and control process), and on olive fruits until the 19th and 16th days of storage in the starter and control process, respectively. S. Enteritidis population showed also a decrease and it was detected until the 21st day of storage in both brine and olive fruits in both cases. The population of L. monocytogenes declined during storage and it was detected until the 31st day of storage in both brine and olive fruits in both cases, showing a longer survival period in comparison to the other two studied pathogens. The presence of the potential probiotic starter did not affect the pathogen survival. The results demonstrated that even though the growth of the pathogenic strains was not supported, they may survive for a long period in a stressful environment of a fermented product with low pH value (4.2) and high salt concentration (6.0%). These results are a valuable contribution to risk assessment studies related to ready to eat foods in general. Copyright © 2013 Elsevier Ltd. All rights reserved.

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

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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.

Effect of caliber size and fat level on the inactivation of E. coli O157:H7 in dry fermented sausages.

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De Souza, James; Ahmed, Rafath; Strange, Philip; Barbut, Shai; Balamurugan, S

2018-02-02

Dry fermented sausages (DFS) have been subject to numerous validation studies, as pathogen reduction heavily relies on both ingredients and processing. In this study the effect of product caliber size (32, 55, 80mm), and fat level (low, 9.67%; high, 18.46% wt/wt) on the inactivation of E. coli O157:H7 during DFS production was examined. Sausages containing a five-strain cocktail of E. coli O157:H7 at 10 7 CFU/g were manufactured and monitored for changes in physicochemical properties and inoculated E. coli O157:H7 numbers were enumerated during the DFS production stages and log reduction rates were calculated. Significant (P0.05) different among sausages of different caliber size or fat levels. No significant (P>0.05) reduction in a w was observed during fermentation of the sausages. However, during the drying phase, sausages with larger caliber sizes required a significantly longer duration of drying to achieve the same a w of smaller caliber size sausages. For instance, to achieve an a w of ≤0.9, following 5days of fermentation/curing, 80mm caliber sausages required up to 27days of drying compared with 13 and 6days for 55 and 32mm caliber size sausages, respectively. Fat levels on the other hand did not significantly (P>0.05) effect the reduction of a w during drying of the sausages. During the fermentation stage there was a significant and rapid reduction in E. coli O157:H7 counts by about 1.1- to 1.4-log units, but was not significantly different among sausages of different caliber size and fat levels. Considering the whole process, only caliber size had a significant effect on log reduction of E. coli O157:H7. ANOVA of log reduction rates of E. coli O157:H7 among sausages of different caliber size and fat levels revealed no significant differences during the fermentation, however, during the drying of the sausages, log reduction rate of E. coli O157:H7 was significantly (PE. coli O157:H7 in high fat large caliber sausages was the lowest at -0.082±0.004 log

Stx1 prophage excision in Escherichia coli strain PA20 confers strong curli and biofilm formation by restoring native mlrA

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Prophage insertions in Escherichia coli O157:H7 mlrA contribute to the low expression of curli fimbriae and biofilm observed in many clinical isolates. Varying levels of CsgD-dependent curli/biofilm expression are restored to strains bearing prophage insertions in mlrA by mutation of regulatory gene...

High-frequency rugose exopolysaccharide production by Vibrio cholerae strains isolated in Haiti.

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Mustafizur Rahman

Full Text Available In October, 2010, epidemic cholera was reported for the first time in Haiti in over 100 years. Establishment of cholera endemicity in Haiti will be dependent in large part on the continued presence of toxigenic V. cholerae O1 in aquatic reservoirs. The rugose phenotype of V. cholerae, characterized by exopolysaccharide production that confers resistance to environmental stress, is a potential contributor to environmental persistence. Using a microbiologic medium promoting high-frequency conversion of smooth to rugose (S-R phenotype, 80 (46.5% of 172 V. cholerae strains isolated from clinical and environmental sources in Haiti were able to convert to a rugose phenotype. Toxigenic V. cholerae O1 strains isolated at the beginning of the epidemic (2010 were significantly less likely to shift to a rugose phenotype than clinical strains isolated in 2012/2013, or environmental strains. Frequency of rugose conversion was influenced by incubation temperature and time. Appearance of the biofilm produced by a Haitian clinical rugose strain (altered biotype El Tor HC16R differed from that of a typical El Tor rugose strain (N16961R by confocal microscopy. On whole-genome SNP analysis, there was no phylogenetic clustering of strains showing an ability to shift to a rugose phenotype. Our data confirm the ability of Haitian clinical (and environmental strains to shift to a protective rugose phenotype, and suggest that factors such as temperature influence the frequency of transition to this phenotype.

Serological characterisation of foot-and-mouth disease type 'O' field isolates from Peru: 1992-1994

International Nuclear Information System (INIS)

Espinoza, A.M.

1998-01-01

Nineteen field isolates of foot-and-mouth disease Virus (FMDV) recovered from bovine epithelial samples corresponding to outbreaks present in different regions of Peru, between 1992-1994 were studied. The relationship of the virus isolates to the O/Urubamba vaccine strain of Peru was determined by the calculation of the 'r' values obtained by the liquid-phase blocking ELISA. All the isolates showed 'r' values higher than 0.66 indicating that the vaccine strain should protect against the field strains. Characterization of the field isolates by a trapping ELISA using a panel of monoclonal antibodies against FMDV O/Switzerland and O/Caseros, showed slight differences in the profiles of the field isolates when compared with the O/Urubamba vaccine strain, but no differences were found among all the isolates. (author)

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

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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

Rapid and Accurate Detection of Bacteriophage Activity against Escherichia coli O157:H7 by Propidium Monoazide Real-Time PCR

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Hui Liu

2014-01-01

Full Text Available Conventional methods to determine the efficacy of bacteriophage (phage for biocontrol of E. coli require several days, due to the need to culture bacteria. Furthermore, cell surface-attached phage particles may lyse bacterial cells during experiments, leading to an overestimation of phage activity. DNA-based real-time quantitative polymerase chain reaction (qPCR is a fast, sensitive, and highly specific means of enumerating pathogens. However, qPCR may underestimate phage activity due to its inability to distinguish viable from nonviable cells. In this study, we evaluated the suitability of propidium monoazide (PMA, a microbial membrane-impermeable dye that inhibits amplification of extracellular DNA and DNA within dead or membrane-compromised cells as a means of using qPCR to identify only intact E. coli cells that survive phage exposure. Escherichia coli O157:H7 strain R508N and 4 phages (T5-like, T1-like, T4-like, and O1-like were studied. Results compared PMA-qPCR and direct plating and confirmed that PMA could successfully inhibit amplification of DNA from compromised/damaged cells E. coli O157:H7. Compared to PMA-qPCR, direct plating overestimated (P < 0.01 phage efficacy as cell surface-attached phage particles lysed E. coli O157:H7 during the plating process. Treatment of samples with PMA in combination with qPCR can therefore be considered beneficial when assessing the efficacy of bacteriophage for biocontrol of E. coli O157:H7.

Functional metagenomics of Escherichia coli O157:H7 interactions with spinach indigenous microorganisms during biofilm formation.

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Michelle Q Carter

Full Text Available The increase in foodborne outbreaks worldwide attributed to fresh fruit and vegetables suggests that produce may serve as an ecological niche for enteric pathogens. Here we examined the interaction of E. coli O157:H7 (EcO157 with spinach leaf indigenous microorganisms during co-colonization and establishment of a mixed biofilm on a stainless steel surface. Stainless steel surface was selected to mimic the surface of produce-processing equipment, where retention of foodborne pathogens such as EcO157 could serve as a potential source for transmission. We observed a positive effect of spinach-associated microbes on the initial attachment of EcO157, but an antagonistic effect on the EcO157 population at the later stage of biofilm formation. Metagenomic analyses of the biofilm community with the GeoChip revealed an extremely diverse community (gene richness, 23409; Shannon-Weiner index H, 9.55. Presence of EcO157 in the mixed biofilm resulted in a significant decrease in the community α-diversity (t test, P<0.05, indicating a putative competition between the pathogen and indigenous spinach microbes. The decrease in the β-diversity of the EcO157-inoculated biofilm at 48 h (ANOVA, P<0.05 suggested a convergent shift in functional composition in response to EcO157 invasion. The success of EcO157 in the mixed biofilm is likely associated with its metabolic potential in utilizing spinach nutrients: the generation time of EcO157 in spinach lysates at 28°C is ~ 38 min, which is comparable to that in rich broth. The significant decrease in the abundance of many genes involved in carbon, nitrogen, and phosphorus cycling in the EcO157-inoculated biofilms (t test, P<0.05 further support our conclusion that competition for essential macronutrients is likely the primary interaction between the EcO157 and indigenous spinach-biofilm species.

Graphene-interfaced electrical biosensor for label-free and sensitive detection of foodborne pathogenic E. coli O157:H7.

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Pandey, Ashish; Gurbuz, Yasar; Ozguz, Volkan; Niazi, Javed H; Qureshi, Anjum

2017-05-15

E. coli O157:H7 is an enterohemorrhagic bacteria responsible for serious foodborne outbreaks that causes diarrhoea, fever and vomiting in humans. Recent foodborne E. coli outbreaks has left a serious concern to public health. Therefore, there is an increasing demand for a simple, rapid and sensitive method for pathogen detection in contaminated foods. In this study, we developed a label-free electrical biosensor interfaced with graphene for sensitive detection of pathogenic bacteria. This biosensor was fabricated by interfacing graphene with interdigitated microelectrodes of capacitors that were biofunctionalized with E. coli O157:H7 specific antibodies for sensitive pathogenic bacteria detection. Here, graphene nanostructures on the sensor surface provided superior chemical properties such as high carrier mobility and biocompatibility with antibodies and bacteria. The sensors transduced the signal based on changes in dielectric properties (capacitance) through (i) polarization of captured cell-surface charges, (ii) cells' internal bioactivity, (iii) cell-wall's electronegativity or dipole moment and their relaxation and (iv) charge carrier mobility of graphene that modulated the electrical properties once the pathogenic E. coli O157:H7 captured on the sensor surface. Sensitive capacitance changes thus observed with graphene based capacitors were specific to E. coli O157:H7 strain with a sensitivity as low as 10-100 cells/ml. The proposed graphene based electrical biosensor provided advantages of speed, sensitivity, specificity and in-situ bacterial detection with no chemical mediators, represents a versatile approach for detection of a wide variety of other pathogens. Copyright © 2016 Elsevier B.V. All rights reserved.

Complete genome sequences of Escherichia coli strains 1303 and ECC-1470 isolated from bovine mastitis

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Leimbach, Andreas; Poehlein, Anja; Witten, Anika; Scheutz, Flemming; Schukken, Ynte|info:eu-repo/dai/nl/075051907; Daniel, Rolf; Dobrindt, Ulrich

2016-01-01

Escherichia coli is the leading causative agent of acute bovine mastitis. Here, we report the complete genome sequence of E. coli O70:H32 strain 1303, isolated from an acute case of bovine mastitis, and E. coli Ont:Hnt strain ECC-1470, isolated from a persistent infection.

Risk of Escherichia coli O157:H7 infection linked to the consumption of beef

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Premarathne, J.M.K.J.K

2017-05-01

Full Text Available Escherichia coli O157:H7 is a major food-borne pathogen that has resulted in numerous outbreaks around the world. Widespread distribution of the organism in various ecological niches impedes the control measures. This study aimed to detect and quantify E. coli O157:H7 in beef sold in wet markets and hypermarkets in Malaysia and to determine the risk of E. coli O157:H7 infection linked to consumption of beef. The rfbO157 and flicH7 primers targeted on somatic antigen (O157 and flagellar antigen (H7 respectively of E. coli O157:H7 was used for the MPN-PCR method. A total of 99 beef samples were collected from local wet markets and hypermarkets. The highest E. coli O157:H7 contamination rate was observed in beef samples collected from wet markets (89.50%, whereas the contamination rate in hyper market A and B were compratively low (35.35 and 20% respectively. However, the microbial load was highest in the beef samples from hypermarket A (1100 MPN/g while E. coli O157:H7 bacterial load in beef samples from hypermarket B and wet market ranged from 3 to 93 MPN/g and 3 to 240 MPN/g, respectively. Using the Quantitative Microbial Risk Assessment (QMRA approach the risk was estimated incorporating the findings of the prevalence study and predictions based on home storage, cooking and consumption patterns. Three different exposure pathways were investigated to estimate the risk associated with contaminated beef and Monte Carlo simulation was used to determine the level of uncertainty. The developed model predicated that consumption of contaminated beef can be accountable for 1.83E+06 E. coli O157:H7 cases per year in Malaysia. The reliability of the model, data gaps and further research needs, is discussed. Through continuous improvement Quantitative Microbial Risk Assessment provides valuable insight into controlling and prevention strategies.

Verocytotoxin producing E. Coli O157 on farms : prevalences, risk factors and transmission

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Schouten, J.M.

2005-01-01

Infection with verocytotoxin producing Escherichia coli (VTEC) O157 in humans can lead to mild or bloody diarrhoea, with e.g. the haemolytic uraemic syndrome (HUS) as possible complication. Cattle appear to be important reservoirs of O157 VTEC. The main

Fate of Escherichia coli O157:H7 (ECOH) in blade tenderized beef prime rib following searing, cooking and holding under commercial conditions

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Undercooked non-intact beef has caused a number of illnesses due to contamination with serotype O157:H7 strains of Escherichia coli (ECOH). Few studies have quantified translocation and/or thermal inactivation of ECOH directly in blade tenderized beef. There have been no such studies for prime rib,...

Survival of Escherichia coli O157:H7 in synthetic gastric fluid after cold and acid habituation in apple juice or trypticase soy broth acidified with hydrochloric acid or organic acids.

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Uljas, H E; Ingham, S C

1998-08-01

Extreme acid tolerance of Escherichia coli O157:H7 has raised doubts about the safety of acidic foods. This study examined whether prior storage in acidic and/or cold conditions enhanced survival of E. coli O157:H7 in synthetic gastric fluid (SGF). Three E. coli O157:H7 strains were stored in trypticase soy broth (TSB; acidified with HCl, malic acid, citric acid, or lactic acid) or pH 3.5 and 6.5 (nonacidic control) apple juice at 4 and 21 degrees C for acids, suggesting that juice constituents other than organic acids protect E. coli O157:H7. Refrigeration combined with low pH best protected cells in apple juice and acidified TSB, but, compared to the nonacidic control, only acidified TSB enhanced subsequent survival in pH 2.5 SGF. Equal survival in SGF occurred after storage in pH 3.5 or 6.5 apple juice at 4 degrees C, suggesting that low temperature alone in apple juice enhanced acid tolerance. Two strains stored at 4 degrees C in TSB containing malic or citric acid subsequently survived better in SGF than cells stored in nonacidified TSB but poorer than cells stored in the presence of HCl. These differences reflect the higher pKa of these organic acids. However, subsequent survival of these strains in SGF was poorer after refrigerated storage in apple juice than in TSB containing citric or malic acids. Cells stored in lactic acid were most likely to be completely eliminated upon transfer to SGF. Differences in survival in storage media or SGF related to strain, storage conditions, or acidifier were consistent and often statistically significant (P acidic beverages may not be affected by the type of acidifier used, the subsequent survival in SGF of this pathogen may be critically dependent on this factor.

Transfer, attachment, and formation of biofilms by Escherichia coli O157:H7 on meat-contact surface materials.

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Simpson Beauchamp, Catherine; Dourou, Dimitra; Geornaras, Ifigenia; Yoon, Yohan; Scanga, John A; Belk, Keith E; Smith, Gary C; Nychas, George-John E; Sofos, John N

2012-06-01

Studies examined the effects of meat-contact material types, inoculation substrate, presence of air at the liquid-solid surface interface during incubation, and incubation substrate on the attachment/transfer and subsequent biofilm formation by Escherichia coli O157:H7 on beef carcass fabrication surface materials. Materials studied as 2 × 5 cm coupons included stainless steel, acetal, polypropylene, and high-density polyethylene. A 6-strain rifampicin-resistant E. coli O157:H7 composite was used to inoculate (6 log CFU/mL, g, or cm²) tryptic soy broth (TSB), beef fat/lean tissue homogenate (FLH), conveyor belt-runoff fluids, ground beef, or beef fat. Coupons of each material were submerged (4 °C, 30 min) in the inoculated fluids or ground beef, or placed between 2 pieces of inoculated beef fat with pressure (20 kg) applied. Attachment/transfer of the pathogen was surface material and substrate dependent, although beef fat appeared to negate differences among surface materials. Beef fat was the most effective (P transfer and subsequent biofilm formation by E. coli O157:H7. The results highlight the importance of thoroughly cleaning soiled surfaces to remove all remnants of beef fat or other organic material that may harbor or protect microbial contaminants during otherwise lethal antimicrobial interventions. © 2012 Institute of Food Technologists®

Inactivation of Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella typhimurium with compounds available in households.

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Yang, Hua; Kendall, Patricia A; Medeiros, Lydia; Sofos, John N

2009-06-01

Solutions of selected household products were tested for their effectiveness against Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium. Hydrogen peroxide (1.5 and 3%), vinegar (2.5 and 5% acetic acid), baking soda (11, 33, and 50% sodium bicarbonate), household bleach (0.0314, 0.0933, and 0.670% sodium hypochlorite), 5% acetic acid (prepared from glacial acetic acid), and 5% citric acid solutions were tested against the three pathogens individually (five-strain composites of each, 10(8) CFU/ml) by using a modified AOAC International suspension test at initial temperatures of 25 and 55degrees C for 1 and 10 min. All bleach solutions (pH 8.36 to 10.14) produced a >5-log reduction of all pathogens tested after 1 min at 25 degrees C, whereas all baking soda solutions (pH 7.32 to 7.55) were ineffective (5-log reduction of both Salmonella Typhimurium and E. coli O157:H7, whereas undiluted vinegar (pH 2.58) had a similar effect only against Salmonella Typhimurium. Compared with 1 min at 25 degrees C, greater reductions of L. monocytogenes (P 3% hydrogen peroxide > undiluted vinegar and 5% acetic acid > 5% citric acid > baking soda (50% sodium bicarbonate). The sensitivity of the tested pathogens to all tested household compounds followed the sequence of Salmonella Typhimurium > E. coli O157: H7 > L. monocytogenes.

Antibacterial Properties of Endophytic Bacteria Isolated from a Fern Species Equisetum arvense L. Against Foodborne Pathogenic Bacteria Staphylococcus aureus and Escherichia coli O157:H7.

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Das, Gitishree; Patra, Jayanta Kumar; Baek, Kwang-Hyun

2017-01-01

Endophytic bacteria (EB) are a rich source of secondary metabolites with medicinal importance. In this study, EB were isolated from the bottle brush herb Equisetum arvense and identified based on 16S rRNA sequencing. Evaluation of its antibacterial potential was conducted using two common foodborne pathogenic bacteria, Staphylococcus aureus ATCC 12600 and Escherichia coli O157:H7 ATCC 43890. Out of 103 identified EB, three species, Streptomyces albolongus, Dermacoccus sp., and Mycobacterium sp., showed significant antibacterial activity against S. aureus with inhibition zones of 45.34 ± 0.15, 43.28 ± 0.19, and 22.98 ± 0.18 mm, respectively, whereas only two species, Streptomyces griseoaurantiacus (EAL196) and Paenibacillus sp. (EAS116), showed moderate antibacterial activity against E. coli O157:H7 with inhibition zones of 9.41 ± 0.29 and 10.44 ± 0.31 mm, respectively. Furthermore, ethyl acetate extract of S. albolongus, Mycobacterium sp., and Dermacoccus sp. showed antibacterial activity against S. aureus, with inhibition zones of 23.43 ± 0.21, 21.18 ± 0.22, and 19.72 ± 0.10 mm, respectively. The methanol extract of Dermacoccus sp. and Paenibacillus sp. showed antibacterial activity against S. aureus and E. coli O157:H7, with inhibition zones of 11.30 ± 0.17 and 10.01 ± 0.21 mm, respectively. Scanning electron microscopy indicated swollen and lysed cell membranes of pathogens treated with ethyl acetate extract. A possible reason might be, likely due to EB metabolites penetrating the bacterial cell membranes and affecting various metabolic functions resulting in lysis. To the best of our knowledge, this is the first study to report that EB from E. arvense can be used as a source of natural antibacterial compounds against foodborne pathogenic bacteria.

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.

Occurrence of Coliform and Escherichia coli Contamination and Absence of Escherichia coli O157:H7 on Romaine Lettuce from Retail Stores in the Upper Midwest.

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Greve, Josephine D; Zietlow, Mark S; Miller, Kevin M; Ellingson, Jay L E

2015-09-01

A total of 720 whole, romaine lettuce heads were purchased from retail locations in the Upper Midwest and assessed for coliform and Escherichia coli contamination and for the presence of E. coli O157:H7. During a 16-month period (August 2010 through December 2011), coliform and E. coli counts were enumerated on Petrifilm, and the presence of E. coli O157:H7 and the virulence gene eae was evaluated by real-time PCR (qPCR). Over half (400 of 720) of the lettuce samples were processed with an immunomagnetic separation step before the qPCR assay. All retail lettuce samples were negative for E. coli O157:H7 when tested with the R.A.P.I.D. LT qPCR targeting a region of the O-antigen, and only two (0.28%) were positive for the eae gene when tested with LightCycler qPCR. On Petrifilm, coliform counts of most lettuce samples (96.4%) were between lettuce samples (98.2%) were lettuce heads. These results contribute to the limited recorded data and understanding of microbial contamination of whole romaine lettuce heads purchased from retail locations, specifically revealing the absence of E. coli O157:H7 and low levels of contamination with coliforms and other E. coli strains.

Antibacterial effect of lactoferricin B on Escherichia coli O157:H7 in ground beef.

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Venkitanarayanan, K S; Zhao, T; Doyle, M P

1999-07-01

The antibacterial activity of lactoferricin B on enterohemorrhagic Escherichia coli O157:H7 in 1% peptone medium and ground beef was studied at 4 and 10 degrees C. In 1% peptone medium, 50 and 100 microg of lactoferricin B per ml reduced E. coli O157:H7 populations by approximately 0.7 and 2.0 log CFU/ml, respectively. Studies comparing the antibacterial effect of lactoferricin B on E. coli O157:H7 in 1% peptone at pH 5.5 and 7.2 did not reveal any significant difference (P > 0.5) at the two pH values. Lactoferricin B (100 microg/g) reduced E. coli O157:H7 population in ground beef by about 0.8 log CFU/g (P 0.5) was observed in the total plate count between treatment and control ground beef samples stored at 4 and 10 degrees C. The antibacterial effect of lactoferricin B on E. coli O157:H7 observed in this study is not of sufficient magnitude to merit its use in ground beef for controlling the pathogen.

Functional Metagenomics of Escherichia coli O157:H7 Interactions with Spinach Indigenous Microorganisms during Biofilm Formation

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Carter, Michelle Q.; Xue, Kai; Brandl, Maria T.; Liu, Feifei; Wu, Liyou; Louie, Jacqueline W.; Mandrell, Robert E.; Zhou, Jizhong

2012-01-01

The increase in foodborne outbreaks worldwide attributed to fresh fruit and vegetables suggests that produce may serve as an ecological niche for enteric pathogens. Here we examined the interaction of E. coli O157:H7 (EcO157) with spinach leaf indigenous microorganisms during co-colonization and establishment of a mixed biofilm on a stainless steel surface. Stainless steel surface was selected to mimic the surface of produce-processing equipment, where retention of foodborne pathogens such as EcO157 could serve as a potential source for transmission. We observed a positive effect of spinach-associated microbes on the initial attachment of EcO157, but an antagonistic effect on the EcO157 population at the later stage of biofilm formation. Metagenomic analyses of the biofilm community with the GeoChip revealed an extremely diverse community (gene richness, 23409; Shannon-Weiner index H, 9.55). Presence of EcO157 in the mixed biofilm resulted in a significant decrease in the community α-diversity (t test, Pbiofilm at 48 h (ANOVA, Pbiofilm is likely associated with its metabolic potential in utilizing spinach nutrients: the generation time of EcO157 in spinach lysates at 28°C is ∼ 38 min, which is comparable to that in rich broth. The significant decrease in the abundance of many genes involved in carbon, nitrogen, and phosphorus cycling in the EcO157-inoculated biofilms (t test, Pbiofilm species. PMID:22957052

Determination of Antimicrobial Activity of Sorrel (Hibiscus sabdariffa) on Esherichia coli O157:H7 Isolated from Food, Veterinary, and Clinical Samples

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Fullerton, Marjorie; Khatiwada, Janak; Johnson, Jacqueline U.; Davis, Shurrita

2011-01-01

Abstract The use of medicinal plants as natural antimicrobial agents is gaining popularity. Sorrel (Hibiscus sabdariffa) is widely used for the treatment of diseases. The objective of this study was to investigate the antimicrobial activity of sorrel on Escherichia coli O157:H7 isolates from food, veterinary, and clinical samples. Phenolics of the calyces were extracted from 10 g of ground, freeze-dried samples using 100 mL of 80% aqueous methanol. Concentrations of 10%, 5%, and 2.5% methanol extract of sorrel were investigated for its antimicrobial activity. Inhibition zones were indicated by a lack of microbial growth due to inhibitory concentrations of sorrel diffused into semisolid culture medium beneath the sorrel-impregnated disk. The results of this experiment showed that the most potent sorrel concentration was 10%, then 5%, and finally 2.5%. The overall mean zone of inhibition for the sorrel extract was 12.66 mm for 10%, 10.75 mm for 5%, and 8.9 mm for 2.5%. The highest inhibition zones (11.16 mm) were observed in veterinary samples, and the lowest (10.57 mm) in the food samples. There were significant (P<.05) differences among mean zones of inhibition found in the food, veterinary, and clinical sources. Based on the source of samples and concentration of sorrel extract, the lowest mean inhibition was 7.00±0.04 mm from clinical samples, and the highest was 15.37±0.61 mm from a food source. These findings indicated that sorrel was effective at all levels in inhibiting E. coli O157:H7; thus it possesses antimicrobial activity and hold great promise as an antimicrobial agent. PMID:21548802

Persistence of Pathogenic and Non-Pathogenic Escherichia coli Strains in Various Tropical Agricultural Soils of India.

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S Naganandhini

Full Text Available The persistence of Shiga-like toxin producing E. coli (STEC strains in the agricultural soil creates serious threat to human health through fresh vegetables growing on them. However, the survival of STEC strains in Indian tropical soils is not yet understood thoroughly. Additionally how the survival of STEC strain in soil diverges with non-pathogenic and genetically modified E. coli strains is also not yet assessed. Hence in the present study, the survival pattern of STEC strain (O157-TNAU was compared with non-pathogenic (MTCC433 and genetically modified (DH5α strains on different tropical agricultural soils and on a vegetable growing medium, cocopeat under controlled condition. The survival pattern clearly discriminated DH5α from MTCC433 and O157-TNAU, which had shorter life (40 days than those compared (60 days. Similarly, among the soils assessed, the red laterite and tropical latosol supported longer survival of O157-TNAU and MTCC433 as compared to wetland and black cotton soils. In cocopeat, O157 recorded significantly longer survival than other two strains. The survival data were successfully analyzed using Double-Weibull model and the modeling parameters were correlated with soil physico-chemical and biological properties using principal component analysis (PCA. The PCA of all the three strains revealed that pH, microbial biomass carbon, dehydrogenase activity and available N and P contents of the soil decided the survival of E. coli strains in those soils and cocopeat. The present research work suggests that the survival of O157 differs in tropical Indian soils due to varied physico-chemical and biological properties and the survival is much shorter than those reported in temperate soils. As the survival pattern of non-pathogenic strain, MTCC433 is similar to O157-TNAU in tropical soils, the former can be used as safe model organism for open field studies.

Reducing of escherichia coli O 157 serotype and cohabitant flora by irradiation in minced meat

International Nuclear Information System (INIS)

Halkman, A.K.; Dogan, H.B.; Yazici, N.

2001-01-01

Escherichia coli O157:H7 was conclusively identified as a pathogen in 1982 following its association with two food-related outbreaks of an unusual gastrointestinal illness. The infectious dose of E. coli O157 is very low, and as a result the organism can be transmitted efficiently not only via contaminated foods but also person to person (Doyle 1991, Karch et al. 1996). Although not definitely linked, consumption of undercooked meats and mainly hamburgers has been strongly implicated in hemorrhagic uremic colitis and hemolytic uremic syndrome. Water and unpasteurized milk are also recognized as sources of outbreaks (Yu and Bruno 1996 , Venkateswaran et al. 1997). Meat and milk products are the most important foods for E. coli O157:H7 outbreaks. Apple cider, drinking and swimming waters are also important for outbreaks. Literature reveals that E. coli O157:H7 is not resistant to the application of radiations. Gamma rays obtained from ''6''0Co ve ''1''3''7Gs sources find wide application in the food protection as these rays eliminate various pathogen including E.coli O157:H7 in the solid foods. Irradiation of food is less effective at temperature below freezing point. In USA beef is allowed to prevent E.coli O157:H7 infection (Farkas et al.1998;Fujikawa et al.1992; Harewood et al.1994;Park et al.1999)

Selection of antibiotics in detection procedure of Escherichia coli O157:H7 in vegetables

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Hoang, Hoang A.; Nhung, Nguyen T. T.

2017-09-01

Detection of Escherichia coli O157:H7 in ready-to-eat fresh vegetables is important since this bacteria is considered as one of the most important pathogens in relation to public health. However, it could be a big challenge for detection of initial low concentrations of E. coli O157:H7 in the samples. In this study, selection of antibiotics that suppress growth of background bacteria to enable detection of E. coli O157:H7 in ready-to-eat fresh vegetables was investigated. Firstly, different combinations of two antibiotics, i.e. novobiocin (N) and vancomycin (V), in BHI broth were conducted. The three antibiotic combinations were preliminary examined their effect on the growth of E. coli O157:H7 and Bacillus spp. in broth based on OD600nm measurement. The combination of both the antibiotics was selected to examine their possibility to support detection of E. coli O157:H7 in vegetables. It was successful when two antibiotics showed their support in detection of E. coli O157:H7 at very low concentration of 2 CFU per one gram of lettuce. Usage of these antibiotics is simple and cheap in the detection procedure and could be applied to other types of ready-to-eat fresh vegetables popular in Vietnam.

Reduction of Escherichia coli O157:H7 viability on leafy green vegetables by treatment with a bacteriophage mixture and trans-cinnamaldehyde.

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Viazis, Stelios; Akhtar, Mastura; Feirtag, Joellen; Diez-Gonzalez, Francisco

2011-02-01

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 has been recognized as a major foodborne pathogen responsible for frequent gastroenteritis outbreaks. Phages and essential oils can be used as a natural antimicrobial method to reduce bacterial pathogens from the food supply. The objective of this study was to determine the effect of a bacteriophage cocktail, BEC8, alone and in combination with the essential oil trans-cinnameldehyde (TC) on the viability of a mixture of EHEC O157:H7 strains applied on whole baby romaine lettuce and baby spinach leaves. The EHEC O157:H7 strains used were Nal(R) mutants of EK27, ATCC 43895, and 472. Exponentially growing cells from tryptic soy (TS) broth cultures were spot inoculated on leaves and dried. EHEC cells were placed at low, medium, and high inoculum levels (10(4), 10(5), and 10(6) CFU/mL, respectively). Appropriate controls, BEC8 (approx. 10(6) PFU/leaf), and TC (0.5% v/v) were applied on treated leaves. The leaves were incubated at 4, 8, 23, and 37 °C in Petri dishes with moistened filter papers. EHEC survival was determined using standard plate count on nalidixic acid (50 μg/mL) Sorbitol MacConkey agar. No survivors were detected when both leaves were treated with BEC8 or TC individually at low inoculum levels after 24 h at 23 and 37 °C. When the EHEC inoculum size increased and/or incubation temperature decreased, the efficacy of BEC8 and TC decreased. However, when the two treatments were combined, no survivors were detected after 10 min at all temperatures and inoculum levels on both leafy greens. These results indicated that the BEC8/TC combination was highly effective against EHEC on both leafy greens. This combination could potentially be used as an antimicrobial to inactivate EHEC O157:H7 and reduce their incidence in the food chain. Copyright © 2010 Elsevier Ltd. All rights reserved.

Earthworms as vectors of Escherichia coli O157:H7 in soil and vermicomposts.

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Williams, A Prysor; Roberts, Paula; Avery, Lisa M; Killham, Ken; Jones, David L

2006-10-01

Survival and movement of Escherichia coli O157:H7 in both soil and vermicompost is of concern with regards to human health. Whilst it is accepted that E. coli O157:H7 can persist for considerable periods in soils, it is not expected to survive thermophilic composting processes. However, the natural behavior of earthworms is increasingly utilized for composting (vermicomposting), and the extent to which earthworms promote the survival and dispersal of the bacterium within such systems is unknown. The faecal material produced by earthworms provides a ready supply of labile organic substrates to surrounding microbes within soil and compost, thus promoting microbial activity. Earthworms can also cause significant movement of organisms through the channels they form. Survival and dispersal of E. coli O157:H7 were monitored in contaminated soil and farmyard manure subjected to earthworm digestion over 21 days. Our findings lead to the conclusion that anecic earthworms such as Lumbricus terrestris may significantly aid vertical movement of E. coli O157 in soil, whereas epigeic earthworms such as Dendrobaena veneta significantly aid lateral movement within compost. Although the presence of earthworms in soil and compost may aid proliferation of E. coli O157 in early stages of contamination, long-term persistence of the pathogen appears to be unaffected.

Inactivation of different strains of Escherichia coli O157:H7 in various apple ciders treated with dimethyl dicarbonate (DMDC) and sulfur dioxide (SO2) as an alternative method.

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Basaran-Akgul, N; Churey, J J; Basaran, P; Worobo, R W

2009-02-01

Escherichia coli has been identified as the causative agent in numerous foodborne illness outbreaks associated with the consumption of fresh apple cider. Apple cider has a pH which is normally below 4.0 and would not be considered a medium capable of supporting the growth of foodborne pathogens. The association of unpasteurized apple cider with foodborne illness due to E. coli O157:H7 has however, led to increased interest in potential alternative methods to produce pathogen free cider. Apple cider was prepared from eight different apple cultivars, inoculated with approximately 10(6)-10(7) CFU of three strains of E. coli O157:H7 per ml (933, ATCC 43889, and ATCC 43895) and tested to determine the effectiveness of sulfur dioxide (SO(2)) and dimethyl dicarbonate (DMDC). Bacterial populations for treated and untreated samples were then enumerated by using non-selective media. Eight different ciders were treated with DMDC (125 and 250 ppm) and SO(2) (25, 50, 75, 100 ppm). Greater than a 5-log reduction was achieved at room temperature with 250 ppm of DMDC and 50 ppm of SO(2) after the incubation time of 6h and 24h, respectively. Addition of DMDC and/or SO(2) may offer an inexpensive alternative to thermal pasteurization for the production of safe apple cider for small apple cider producers.

Host cell interactions of outer membrane vesicle-associated virulence factors of enterohemorrhagic Escherichia coli O157: Intracellular delivery, trafficking and mechanisms of cell injury

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Greune, Lilo; Jarosch, Kevin-André; Steil, Daniel; Zhang, Wenlan; He, Xiaohua; Lloubes, Roland; Fruth, Angelika; Kim, Kwang Sik; Schmidt, M. Alexander; Dobrindt, Ulrich; Mellmann, Alexander; Karch, Helge

2017-01-01

Outer membrane vesicles (OMVs) are important tools in bacterial virulence but their role in the pathogenesis of infections caused by enterohemorrhagic Escherichia coli (EHEC) O157, the leading cause of life-threatening hemolytic uremic syndrome, is poorly understood. Using proteomics, electron and confocal laser scanning microscopy, immunoblotting, and bioassays, we investigated OMVs secreted by EHEC O157 clinical isolates for virulence factors cargoes, interactions with pathogenetically relevant human cells, and mechanisms of cell injury. We demonstrate that O157 OMVs carry a cocktail of key virulence factors of EHEC O157 including Shiga toxin 2a (Stx2a), cytolethal distending toxin V (CdtV), EHEC hemolysin, and flagellin. The toxins are internalized by cells via dynamin-dependent endocytosis of OMVs and differentially separate from vesicles during intracellular trafficking. Stx2a and CdtV-B, the DNase-like CdtV subunit, separate from OMVs in early endosomes. Stx2a is trafficked, in association with its receptor globotriaosylceramide within detergent-resistant membranes, to the Golgi complex and the endoplasmic reticulum from where the catalytic Stx2a A1 fragment is translocated to the cytosol. CdtV-B is, after its retrograde transport to the endoplasmic reticulum, translocated to the nucleus to reach DNA. CdtV-A and CdtV-C subunits remain OMV-associated and are sorted with OMVs to lysosomes. EHEC hemolysin separates from OMVs in lysosomes and targets mitochondria. The OMV-delivered CdtV-B causes cellular DNA damage, which activates DNA damage responses leading to G2 cell cycle arrest. The arrested cells ultimately die of apoptosis induced by Stx2a and CdtV via caspase-9 activation. By demonstrating that naturally secreted EHEC O157 OMVs carry and deliver into cells a cocktail of biologically active virulence factors, thereby causing cell death, and by performing first comprehensive analysis of intracellular trafficking of OMVs and OMV-delivered virulence factors

Leakage of Intracellular UV Materials of High Hydrostatic Pressure-Injured Escherichia Coli O157:H7 Strains in Tomato Juice

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The effect of high hydrostatic pressure (HHP) treatment on inactivation, injury and recovery of Salmonella Enteritidis and Escherichia coli O157:H7 cocktail inoculated in tomato juice (pH 4.1) and phosphate buffer saline (PBS. pH 7.2) at 8.0 log CFU/ml and treated at 350, 400, 450 MPa for 20 min at ...

75 FR 10460 - Improving Tracing Procedures for E. coli O157:H7 Positive Raw Beef Product

Science.gov (United States)

2010-03-08

... Tracing Procedures for E. coli O157:H7 Positive Raw Beef Product AGENCY: Food Safety and Inspection... has found positive for Escherichia coli (E. coli) O157:H7. FSIS will also discuss additional verification activities the Agency will conduct at suppliers in response to positive E. coli O157:H7 results...

Plant lesions promote the rapid multiplication of Escherichia coli O157:H7 on post-harvest lettuce

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Several outbreaks of Escherichia coli O157:H7 (EcO157) infections have been associated with minimally processed leafy vegetables in the U.S. Harvesting and processing cause plant tissue damage. In order to assess the role of plant tissue damage in the contamination of leafy greens with EcO157, the e...

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

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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.

Complete Genome Sequences of Two Escherichia coli O145:H28 Outbreak Strains of Food Origin

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Cooper, Kerry K.; Mandrell, Robert E.; Louie, Jacqueline W.; Korlach, Jonas; Clark, Tyson A.; Parker, Craig T.; Huynh, Steven; Chain, Patrick S. G.; Ahmed, Sanaa; Carter, Michelle Qiu

2014-01-01

Escherichia coli O145:H28 strain RM12581 was isolated from bagged romaine lettuce during a 2010 U.S. lettuce-associated outbreak. E. coli O145:H28 strain RM12761 was isolated from ice cream during a 2007 ice cream-associated outbreak in Belgium. Here we report the complete genome sequences and annotation of both strains.

Synthesis of O-serogroup specific positive controls and real-time PCR standards for nine clinically relevant non-O157 STECs.

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Conrad, Cheyenne C; Gilroyed, Brandon H; McAllister, Tim A; Reuter, Tim

2012-10-01

Non-O157 Shiga toxin producing Escherichia coli (STEC) are gaining recognition as human pathogens, but no standardized method exists to identify them. Sequence analysis revealed that STEC can be classified on the base of variable O antigen regions into different O serotypes. Polymerase chain reaction is a powerful technique for thorough screening and complex diagnosis for these pathogens, but requires a positive control to verify qualitative and/or quantitative DNA-fragment amplification. Due to the pathogenic nature of STEC, controls are not readily available and cell culturing of STEC reference strains requires biosafety conditions of level 2 or higher. In order to bypass this limitation, controls of stacked O-type specific DNA-fragments coding for primer recognition sites were designed to screen for nine STEC serotypes frequently associated with human infection. The synthetic controls were amplified by PCR, cloned into a plasmid vector and transferred into bacteria host cells. Plasmids amplified by bacterial expression were purified, serially diluted and tested as standards for real-time PCR using SYBR Green and TaqMan assays. Utility of synthetic DNA controls was demonstrated in conventional and real-time PCR assays and validated with DNA from natural STEC strains. Copyright © 2012 Elsevier B.V. All rights reserved.

Fate of Escherichia coli O157: H7 in agricultural soils amended with different organic fertilizers.

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Yao, Zhiyuan; Yang, Li; Wang, Haizhen; Wu, Jianjun; Xu, Jianming

2015-10-15

Five organic fertilizers (vermicompost, pig manure, chicken manure, peat and oil residue) were applied to agricultural soils to study their effects on the survival of Escherichia coli O157:H7 (E. coli O157:H7). Results showed that E. coli O157:H7 survival changed greatly after organic fertilizers application, with shorter td values (survival time needed to reach the detection limit of 100 CFU g(-1)) (12.57±6.57 days) in soils amended with chicken manure and the longest (25.65±7.12 days) in soils amended with pig manure. Soil pH, EC and free Fe/Al (hydro) oxides were significant explanatory factors for E. coli O157:H7 survival in the original soils. Soil constituents (minerals and organic matter) and changes in their surface charges with pH increased the effect of soil pH on E. coli O157:H7 survival. However, electrical conductivity played a more important role in regulating E. coli O157:H7 survival in fertilizer-amended soils. This study highlighted the importance of choosing appropriate organic fertilizers in the preharvest environment to reduce food-borne bacterial contamination. Copyright © 2015 Elsevier B.V. All rights reserved.

Isolation of non-sulphur photosynthetic bacterial strains efficient in hydrogen production at elevated temperatures

Energy Technology Data Exchange (ETDEWEB)

Singh, S.P.; Srivastava, S.C. (Banaras Hindu Univ., Varanasi (IN). Centre of Advanced Study in Botany)

1991-01-01

Four strains of non-sulphur photosynthetic bacteria were isolated from root zone associations of aquatic plants like Azolla, Salvinia and Eichhornia, as well as the deep-water rice. Based on the gross cell morphology and pigmentation, the isolates resembled Rhodopseudomonas sp. and have been designated as BHU strains 1 to 4, respectively. When subjected to elevated temperature (from 33-45{sup o}C), substantial growth/hydrogen production could be observed only in strains 1 and 4. Strains 2 and 3 on the other hand, showed diminished growth and negligible hydrogen photoproduction. The BHU strains 1 and 4 have been selected as the most active (thermostable) hydrogen producing strains of local origin as far as the Indian tropical climate is concerned. (author).

Escherichia coli O157:H7 outbreak associated with consumption of ground beef, June-July 2002.

Science.gov (United States)

Vogt, Richard L; Dippold, Laura

2005-01-01

A case-control and environmental study tested the hypothesis that purchasing and eating ground beef from a specific source was the cause of a cluster of cases of hemolytic uremic syndrome (HUS) and Escherichia coli (E. coli) O157:H7 gastroenteritis. A case-control study comparing risk factors was conducted over the telephone on nine case-patients with 23 selected controls. An environmental investigation was conducted that consisted of reviewing beef handling practices at a specific local supermarket and obtaining ground beef samples from the store and two households with case-patients. The analysis of the case-control study showed that eight case-patients (89%) purchased ground beef at Grocery Chain A compared with four controls who did not develop illness (17%) (matched odds ratio=undefined; 95% confidence interval 2.8, infinity; p=0.006). The environmental investigation showed that Grocery Chain A received meat from Meatpacker A. Laboratory analysis of meat samples from Meatpacker A and Grocery Chain A and stool samples from some patients recovered an identical strain of E. coli O157:H7 according to pulse-field gel electrophoresis. Both the case-control and environmental studies showed that purchasing ground beef at Grocery Chain A, which received ground beef from Meatpacker A, was the major risk factor for illness in eight case-patients; the ninth case-patient was found to be unrelated to the outbreak. Furthermore, meat from Meatpacker A was associated with a nationwide outbreak of E. coli O157:H7 illness that resulted in the second largest recall of beef in U.S. history at the time.

Evaluation of Lactobacillus strains for selected probiotic properties.

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Turková, Kristýna; Mavrič, Anja; Narat, Mojca; Rittich, Bohuslav; Spanová, Alena; Rogelj, Irena; Matijašić, Bojana Bogovič

2013-07-01

Eleven strains of Lactobacillus collected in the Culture Collection of Dairy Microorganisms (CCDM) were evaluated for selected probiotic properties such as survival in gastrointestinal fluids, antimicrobial activity, and competition with non-toxigenic Escherichia coli O157:H7 for adhesion on Caco-2 cells. The viable count of lactobacilli was reduced during 3-h incubation in gastric fluid followed by 3-h incubation in intestinal fluid. All strains showed antimicrobial activity and the three most effective strains inhibited the growth of at least 16 indicator strains. Antimicrobial metabolites of seven strains active against Lactobacillus and Clostridium indicator strains were found to be sensitive to proteinase K and trypsin, which indicates their proteinaceous nature. The degree of competitive inhibition of non-toxigenic E. coli O157:H7 adhesion on the surface of Caco-2 cells was strain-dependent. A significant decrease (P strains were selected for additional studies of antimicrobial activity, i.e., Lactobacillus gasseri CCDM 215, Lactobacillus acidophilus CCDM 149, and Lactobacillus helveticus CCDM 82.

High efficiency generalized transduction in Escherichia coli O157:H7 [v1; ref status: indexed, http://f1000r.es/8f

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Martin G Marinus

2013-01-01

Full Text Available Genetic manipulation in enterohemorrhagic E. coli O157:H7 is currently restricted to recombineering, a method that utilizes the recombination system of bacteriophage lambda, to introduce gene replacements and base changes inter alia into the genome. Bacteriophage 933W is a prophage in E. coli O157:H7 strain EDL933, which encodes the genes (stx2AB for the production of Shiga toxin which is the basis for the potentially fatal Hemolytic Uremic Syndrome in infected humans. We replaced the stx2AB genes with a kanamycin cassette using recombineering. After induction of the prophage by ultra-violet light, we found that bacteriophage lysates were capable of transducing to wildtype, point mutations in the lactose, arabinose and maltose genes. The lysates could also transduce tetracycline resistant cassettes. Bacteriophage 933W is also efficient at transducing markers in E. coli K-12. Co-transduction experiments indicated that the maximal amount of transferred DNA was likely the size of the bacteriophage genome, 61 kB. All tested transductants, in both E. coli K-12 and O157:H7, were kanamycin-sensitive indicating that the transducing particles contained host DNA.

Assessing the growth of Escherichia coli O157:H7 and Salmonella in spinach, lettuce, parsley and chard extracts at different storage temperatures.

Science.gov (United States)

Posada-Izquierdo, G; Del Rosal, S; Valero, A; Zurera, G; Sant'Ana, A S; Alvarenga, V O; Pérez-Rodríguez, F

2016-06-01

The objective of this work was to study the growth potential of Escherichia coli O157:H7 and Salmonella spp. in leafy vegetable extracts at different temperature conditions. Cocktails of five strains of E. coli O157:H7 and of Salmonella enterica were used. Inoculated aqueous vegetable extracts were incubated at 8, 10, 16 and 20°C during 21 days. Microbial growth was monitored using Bioscreen C(®) . In spinach extract, results showed that for E. coli O157:H7 and Salmonella significant differences (P parsley showed the lowest values of μabs , below 0·008 h(-1) . The coefficients of variance (CoV) calculated for the different replicates evidenced that at low temperature (8°C) a more variable behaviour of both pathogens is expected (CoV > 180%). This study provides evidence that aqueous extracts from vegetable tissues can result in distinct growth niche producing different response in various types of vegetables. Finally, these results can be used as basis to establish risk rankings of pathogens and leafy vegetable matrices with relation to their potential growth. © 2016 The Society for Applied Microbiology.

[Antibiotic resistance pattern of 24, 526 strains of Vibrio cholerae O1 isolated in Mexico from 1991 to 1993].

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Giono-Cerezo, S; Zárate, A; Gutiérrez, L; Valdespino, J L

1994-01-01

Profile of antimicrobial resistance by Kirby-Bauer method was performed on 24526 Vibrio cholerae O1 strains isolated in México (1991-1993) from fecal swabs in cholera cases and from asymptomatic carriers. Minimal inhibitory concentration (MIC) tests for tetracycline (Te) and doxycycline (D) were done on selected strains. Single antibiotic discs were used at concentrations of: Te, 30 micrograms; D, 30 micrograms; erythromycin (E), 15 micrograms; chloramphenicol (CM), 30 micrograms; ampicillin (AM), 10 micrograms; trimethoprim-sulfamethoxazole (SXT) 1.25 micrograms/23.75 micrograms. Strains whose halos were of a smaller diameter than the intermediate value were considered resistant. It is important to maintain surveillance on antimicrobial susceptibility as epidemiological marker on geographical selected areas in order to detect changes of resistant patterns.

Towards a Molecular Definition of Enterohemorrhagic Escherichia coli (EHEC): Detection of Genes Located on O Island 57 as Markers To Distinguish EHEC from Closely Related Enteropathogenic E. coli Strains

Science.gov (United States)

Delannoy, Sabine; Beutin, Lothar

2013-01-01

Among strains of Shiga-toxin (Stx) producing Escherichia coli (STEC), seven serogroups (O26, O45, O103, O111, O121, O145, and O157) are associated with severe clinical illness in humans. These strains are also called enterohemorrhagic E. coli (EHEC), and the development of methods for their reliable detection from food has been challenging thus far. PCR detection of major EHEC virulence genes stx1, stx2, eae, and O-serogroup-specific genes is useful but does not identify EHEC strains specifically. Searching for the presence of additional genes issued from E. coli O157:H7 genomic islands OI-122 and OI-71 increases the specificity but does not clearly discriminate EHEC from enteropathogenic E. coli (EPEC) strains. Here, we identified two putative genes, called Z2098 and Z2099, from the genomic island OI-57 that were closely associated with EHEC and their stx-negative derivative strains (87% for Z2098 and 91% for Z2099). Z2098 and Z2099 were rarely found in EPEC (10% for Z2098 and 12% for Z2099), STEC (2 and 15%), and apathogenic E. coli (1% each) strains. Our findings indicate that Z2098 and Z2099 are useful genetic markers for a more targeted diagnosis of typical EHEC and new emerging EHEC strains. PMID:23325824

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.

Subtractive Inhibition Assay for the Detection of E. coli O157:H7 Using Surface Plasmon Resonance

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Chengyan Si

2011-03-01

Full Text Available A surface plasmon resonance (SPR immunosensor was developed for the detection of E. coli O157:H7 by means of a new subtractive inhibition assay. In the subtractive inhibition assay, E. coli O157:H7 cells and goat polyclonal antibodies for E. coli O157:H7 were incubated for a short of time, and then the E. coli O157:H7 cells which bound antibodies were removed by a stepwise centrifugation process. The remaining free unbound antibodies were detected through interaction with rabbit anti-goat IgG polyclonal antibodies immobilized on the sensor chip using a BIAcore 3000 biosensor. The results showed that the signal was inversely correlated with the concentration of E. coli O157:H7 cells in a range from 3.0 × 104 to 3.0 × 108 cfu/mL with a detection limit of 3.0 × 104 cfu/mL. Compared with direct SPR by immobilizing antibodies on the chip surface to capture the bacterial cells and ELISA for E. coli O157:H7 (detection limit: both 3.0 × 105 cfu/mL in this paper, the detection limit of subtractive inhibition assay method was reduced by one order of magnitude. The method simplifies bacterial cell detection to protein-protein interaction, which has the potential for providing a practical alternative for the monitoring of E. coli O157:H7 and other pathogens.

Survival of Escherichia coli O157:H7 in ground beef jerky assessed on two plating media.

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Harrison, J A; Harrison, M A; Rose, R A

1998-01-01

Recent outbreaks of food-borne illness due to Salmonella spp. in beef jerky and Escherichia coli O157:H7 in venison jerky, coupled with the fact that a variety of preparation methods and dying procedures abound, raise concern over the safety of processed meat products made in the home. The potential of injured bacterial cells to regain the ability to cause illness is a particular threat with pathogens such as E. coli O157:H7, which is believed to have a low infectious dose. This study examined the efficacy of various methods of jerky preparation in reducing populations of E, coli O157:H7 in ground beef jerky and compared the recovery rate of E. coli O157:H7 on two selective plating media, modified sorbitol MacConkey agar (MSMA) and modified eosin methylene blue agar (MEMB). Populations of E. coli O157:H7 in both heated and unheated samples exhibited a greater decline during drying when a nitrite and salt cure mix was added during jerky preparation. When recovery of E. coli O157:H7 on MSMA and MEMB was compared, a trend toward slightly higher recovery rates with MEMB was observed. On the basis of these results, MEMB is a suitable alternative to MSMA for the recovery of E. coli O157:H7 from heated and dried meat samples similar to beef jerky.

Enterobacterial repetitive intergenic consensus sequences and the PCR to generate fingerprints of genomic DNAs from Vibrio cholerae O1, O139, and non-O1 strains.

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Rivera, I G; Chowdhury, M A; Huq, A; Jacobs, D; Martins, M T; Colwell, R R

1995-08-01

Enterobacterial repetitive intergenic consensus (ERIC) sequence polymorphism was studied in Vibrio Cholerae strains isolated before and after the cholera epidemic in Brazil (in 1991), along with epidemic strains from Peru, Mexico, and India, by PCR. A total of 17 fingerprint patterns (FPs) were detected in the V. cholerae strains examined; 96.7% of the toxigenic V. cholerae O1 strains and 100% of the O139 serogroup strains were found to belong to the same FP group comprising four fragments (FP1). The nontoxigenic V. cholerae O1 also yielded four fragments but constituted a different FP group (FP2). A total of 15 different patterns were observed among the V. cholerae non-O1 strains. Two patterns were observed most frequently for V. cholerae non-01 strains, 25% of which have FP3, with five fragments, and 16.7% of which have FP4, with two fragments. Three fragments, 1.75, 0.79, and 0.5 kb, were found to be common to both toxigenic and nontoxigenic V. cholerae O1 strains as well as to group FP3, containing V. cholerae non-O1 strains. Two fragments of group FP3, 1.3 and 1.0 kb, were present in FP1 and FP2 respectively. The 0.5-kb fragment was common to all strains and serogroups of V. cholerae analyzed. It is concluded from the results of this study, based on DNA FPs of environmental isolates, that it is possible to detect an emerging virulent strain in a cholera-endemic region. ERIC-PCR constitutes a powerful tool for determination of the virulence potential of V. cholerae O1 strains isolated in surveillance programs and for molecular epidemiological investigations.

Photocatalysis-assisted water filtration: using TiO2-coated vertically aligned multi-walled carbon nanotube array for removal of Escherichia coli O157:H7.

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Oza, Goldie; Pandey, Sunil; Gupta, Arvind; Shinde, Sachin; Mewada, Ashmi; Jagadale, Pravin; Sharon, Maheshwar; Sharon, Madhuri

2013-10-01

A porous ceramic was coated with vertically aligned multi-walled carbon nanotubes (MWCNTs) by spray pyrolysis. Titanium dioxide (TiO2) nanoparticles were then coated onto this densely aligned MWCNT. The presence of TiO2/MWCNT interfacial arrays was confirmed by X-ray diffraction (XRD), scanning electron microscope-energy dispersive analysis of X-ray (SEM-EDAX) and transmission electron microscope (TEM). This is a novel report in which water loaded with a most dreadful enterohemorrhagic pathogenic strain of Escherichia coli O157:H7 was filtered through TiO2/MWCNT coated porous ceramic filter and then analysed. Bacterial removal performance was found to be significantly lower in control i.e. plain porous ceramic (Paligned MWCNT network. © 2013 Elsevier B.V. All rights reserved.

Pathogenic strains of Yersinia enterocolitica isolated from domestic dogs (Canis familiaris) belonging to farmers are of the same subtype as pathogenic Y. enterocolitica strains isolated from humans and may be a source of human infection in Jiangsu Province, China.

Science.gov (United States)

Wang, Xin; Cui, Zhigang; Wang, Hua; Tang, Liuying; Yang, Jinchuan; Gu, Ling; Jin, Dong; Luo, Longze; Qiu, Haiyan; Xiao, Yuchun; Xiong, Haiping; Kan, Biao; Xu, Jianguo; Jing, Huaiqi

2010-05-01

We isolated 326 Yersinia enterocolitica strains from 5,919 specimens from patients with diarrhea at outpatient clinics, livestock, poultry, wild animals, insect vectors, food, and the environment in the cities of Nantong and Xuzhou in Jiangsu Province, China, from 2004 to 2008. The results showed that the 12 pathogenic strains were of the O:3 serotype. Six strains were isolated from domestic dogs (Canis familiaris) belonging to farmers and were found to be the primary carriers of pathogenic Y. enterocolitica strains, especially in Xuzhou. Pulsed-field gel electrophoresis analysis of the pathogenic strains from dogs belonging to farmers showed that they shared the same patterns as strains from diarrhea patients isolated in 1994. This indicates that the strains from domestic dogs have a close correlation with the strains causing human infections.

Lytic bacteriophages reduce Escherichia coli O157: H7 on fresh cut lettuce introduced through cross-contamination.

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Ferguson, Sean; Roberts, Cheryl; Handy, Eric; Sharma, Manan

2013-01-01

The role of lytic bacteriophages in preventing cross contamination of produce has not been evaluated. A cocktail of three lytic phages specific for E. coli O157:H7 (EcoShield™) or a control (phosphate buffered saline, PBS) was applied to lettuce by either; (1) immersion of lettuce in 500 ml of EcoShield™ 8.3 log PFU/ml or 9.8 log PFU/ml for up to 2 min before inoculation with E. coli O157:H7; (2) spray-application of EcoShield™ (9.3 log PFU/ml) to lettuce after inoculation with E. coli O157:H7 (4.10 CFU/cm 2 ) following exposure to 50 μg/ml chlorine for 30 sec. After immersion studies, lettuce was spot-inoculated with E. coli O157:H7 (2.38 CFU/cm 2 ). Phage-treated, inoculated lettuce pieces were stored at 4°C for and analyzed for E. coli O157:H7 populations for up to 7 d. Immersion of lettuce in 9.8 log PFU/ml EcoShield™ for 2 min significantly (p PFU/ml) resulted in the deposition of high concentrations (7.8 log log PFU/cm 2 ) of bacteriophages on the surface of fresh cut lettuce, potentially contributing to the efficacy of the lytic phages on lettuce. Spraying phages on to inoculated fresh cut lettuce after being washed in hypochlorite solution was significantly more effective in reducing E. coli O157:H7 populations (2.22 log CFU/cm 2 ) on day 0 compared with control treatments (4.10 log CFU/cm 2 ). Both immersion and spray treatments provided protection from E. coli O157:H7 contamination on lettuce, but spray application of lytic bacteriophages to lettuce was more effective in immediately reducing E. coli O157:H7 populations fresh cut lettuce.

Transfer of Escherichia coli O157:H7 from equipment surfaces to fresh-cut leafy greens during processing in a model pilot-plant production line with sanitizer-free water.

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Buchholz, Annemarie L; Davidson, Gordon R; Marks, Bradley P; Todd, Ewen C D; Ryser, Elliot T

2012-11-01

Escherichia coli O157:H7 contamination of fresh-cut leafy greens has become a public health concern as a result of several large outbreaks. The goal of this study was to generate baseline data for E. coli O157:H7 transfer from product-inoculated equipment surfaces to uninoculated lettuce during pilot-scale processing without a sanitizer. Uninoculated cored heads of iceberg and romaine lettuce (22.7 kg) were processed using a commercial shredder, step conveyor, 3.3-m flume tank with sanitizer-free tap water, shaker table, and centrifugal dryer, followed by 22.7 kg of product that had been dip inoculated to contain ∼10(6), 10(4), or 10(2) CFU/g of a four-strain avirulent, green fluorescent protein-labeled, ampicillin-resistant E. coli O157:H7 cocktail. After draining the flume tank and refilling the holding tank with tap water, 90.8 kg of uninoculated product was similarly processed and collected in ∼5-kg aliquots. After processing, 42 equipment surface samples and 46 iceberg or 36 romaine lettuce samples (25 g each) from the collection baskets were quantitatively examined for E. coli O157:H7 by direct plating or membrane filtration using tryptic soy agar containing 0.6% yeast extract and 100 ppm of ampicillin. Initially, the greatest E. coli O157:H7 transfer was seen from inoculated lettuce to the shredder and conveyor belt, with all equipment surface populations decreasing 90 to 99% after processing 90.8 kg of uncontaminated product. After processing lettuce containing 10(6) or 10(4) E. coli O157:H7 CFU/g followed by uninoculated lettuce, E. coli O157:H7 was quantifiable throughout the entire 90.8 kg of product. At an inoculation level of 10(2) CFU/g, E. coli O157:H7 was consistently detected in the first 21.2 kg of previously uninoculated lettuce at 2 to 3 log CFU/100 g and transferred to 78 kg of product. These baseline E. coli O157:H7 transfer results will help determine the degree of sanitizer efficacy required to better ensure the safety of fresh-cut leafy

Agrochemicals indirectly increase survival of E. coli O157:H7 and indicator bacteria by reducing ecosystem services.

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Staley, Zachery R; Rohr, Jason R; Senkbeil, Jacob K; Harwood, Valerie J

Storm water and agricultural runoff frequently contain agrochemicals, fecal indicator bacteria (FIB), and zoonotic pathogens. Entry of such contaminants into aquatic ecosystems may affect ecology and human health. This study tested the hypothesis that the herbicide atrazine and the fungicide chlorothalonil indirectly affect the survival of FIB (Escherichia coli and Enterococcus faecalis) and a pathogen (E. coli O157:H7) by altering densities of protozoan predators or by altering competition from autochthonous bacteria. Streptomycin-resistant E. coli, En. faecalis, and E. coli O157:H7 were added to microcosms composed of Florida river water containing natural protozoan and bacterial populations. FIB, pathogen, and protozoan densities were monitored over six days. Known metabolic inhibitors, cycloheximide and streptomycin, were used to inhibit autochthonous protozoa or bacteria, respectively. The inhibitors made it possible to isolate the effects of predation or competition on survival of allochthonous bacteria, and each treatment increased the survival of FIB and pathogens. Chlorothalonil's effect was similar to that of cycloheximide, significantly reducing protozoan densities and elevating densities of FIB and pathogens relative to the control. Atrazine treatment did not affect protozoan densities, but, through an effect on competition, resulted in significantly greater densities of En. faecalis and E. coli O157:H7. Hence, by reducing predaceous protozoa and bacterial competitors that facilitate purifying water bodies of FIBs and human pathogens, chlorothalonil and atrazine indirectly diminished an ecosystem service of fresh water.

Modified Vero cell induced by Bifidobacterium bifidum inhibits enterohemorrhagic Escherichia coli O157:H7 cytopathic effect

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Tahamtan, Y.

2014-11-01

Full Text Available Enterohemorrhagic Escherichia coli (EHEC, such as E. coli O157:H7, are emerging food-borne pathogens worldwide. This micro-organism can damage the epithelial tissue of the large intestine. The cytotoxic effects can be neutralized by probiotics such as Bifidobacterium bifidum. Probiotics are viable cells that have beneficial effects on the health of the host. The preventing activity of B. bifidum against E. coli O157 was studied using a Vero cell model. Vero cell was pretreated with viable B. bifidum and incubated for either 3 h to 24 h and then collected from the cell to make modified Vero cell (MVC. Indirect antibacterial effects of B. bifidum were demonstrated by reduction of attachment of E. coli O157:H7 to MVC. The maximum reduction was resulted in pretreatment of Vero cell with B. bifidum for 24 h before infection. B. bifidum attenuated E. coli O157:H7 attachment to MVC up to 10 days of incubation. To our knowledge, MCV prevented Vero cell line injury induced by E. coli O157:H7. Therefore, B. bifidum can be used for inhibition of E. coli O157:H7 cytopathic effect (CPE in Vero cell model, even as pretreatment of the cell line.

Relative nephroprotection during Escherichia coli O157:H7 infections: association with intravenous volume expansion.

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Ake, Julie A; Jelacic, Srdjan; Ciol, Marcia A; Watkins, Sandra L; Murray, Karen F; Christie, Dennis L; Klein, Eileen J; Tarr, Phillip I

2005-06-01

The hemolytic uremic syndrome (HUS) consists of hemolytic anemia, thrombocytopenia, and renal failure. HUS is often precipitated by gastrointestinal infection with Shiga toxin-producing Escherichia coli and is characterized by a variety of prothrombotic host abnormalities. In much of the world, E coli O157:H7 is the major cause of HUS. HUS can be categorized as either oligoanuric (which probably signifies acute tubular necrosis) or nonoligoanuric. Children with oligoanuric renal failure during HUS generally require dialysis, have more complicated courses, and are probably at increased risk for chronic sequelae than are children who experience nonoligoanuric HUS. Oligoanuric HUS should be avoided, if possible. The presentation to medical care of a child with definite or possible E coli O157:H7 infections but before HUS ensues affords a potential opportunity to ameliorate the course of the subsequent renal failure. However, it is not known whether events that occur early in E coli O157:H7 infections, particularly measures to expand circulating volume, affect the likelihood of experiencing oligoanuric HUS if renal failure develops. We attempted to assess whether pre-HUS interventions and events, especially the volume and sodium content of intravenous fluids administered early in illness, affect the risk for developing oligoanuric HUS after E coli O157:H7 infections. We performed a prospective cohort study of 29 children with HUS that was confirmed microbiologically to be caused by E coli O157:H7. Infected children were enrolled when they presented with acute bloody diarrhea or as contacts of patients who were known to be infected with E coli O157:H7, or if they had culture-confirmed infection, or if they presented with HUS. HUS was defined as hemolytic anemia (hematocrit parenteral fluid administered, were recorded and entered into analysis. Estimates of odds ratios were adjusted for possible confounding effects using logistic regression analysis. Twenty-nine children

Effect of CuO Nanoparticles over Isolated Bacterial Strains from Agricultural Soil

International Nuclear Information System (INIS)

Concha-Guerrero, S.I.; Pinon-Castillo, H.A.; Luna-Velasco, A.; Orrantia-Borunda, E.; Brito, E.M.S.; Tarango-Rivero, S.H.; Caretta, C.A.; Duran, R.

2014-01-01

The increased use of the nanoparticles (NPs) on several processes is notorious. In contrast the eco toxicological effects of NPs have been scarcely studied. The main current researches are related to the oxide metallic NPs. In the present work, fifty-six bacterial strains were isolated from soil, comprising 17 different OTUs distributed into 3 classes: Bacilli (36 strains), Flavobacteria (2 strains), and Gamma proteobacteria (18 strains). Copper oxide nanoparticles (CuONPs) were synthesized using a process of chemical precipitation. The obtained CuONPs have a spherical shape and primary size less than 17 nm. Twenty-one strains were used to evaluate the cytotoxicity of CuONPs and 11 of these strains showed high sensibility. Among those 11 strains, 4 (Brevibacillus later osporus strain CSS8, Chryseobacterium indoltheticum strain CSA28, and Pantoea ananatis strains CSA34 and CSA35) were selected to determine the kind of damage produced. The CuONPs toxic effect was observed at expositions over 25 mg·L -1 and the damage to cell membrane above 160 mg·L -1 . The electron microscopy showed the formation of cavities, holes, membrane degradation, blebs, cellular collapse, and lysis. These toxic effects may probably be due to the ions interaction, the oxide-reduction reactions, and the generation of reactive species

Control of foodborne pathogens on fresh-cut fruit by a novel strain of Pseudomonas graminis.

Science.gov (United States)

Alegre, Isabel; Viñas, Inmaculada; Usall, Josep; Teixidó, Neus; Figge, Marian J; Abadias, Maribel

2013-06-01

The consumption of fresh-cut fruit has substantially risen over the last few years, leading to an increase in the number of outbreaks associated with fruit. Moreover, consumers are currently demanding wholesome, fresh-like, safe foods without added chemicals. As a response, the aim of this study was to determine if the naturally occurring microorganisms on fruit are "competitive with" or "antagonistic to" potentially encountered pathogens. Of the 97 and 107 isolates tested by co-inoculation with Escherichia coli O157:H7, Salmonella and Listeria innocua on fresh-cut apple and peach, respectively, and stored at 20 °C, seven showed a strong antagonistic capacity (more than 1-log unit reduction). One of the isolates, CPA-7, achieved the best reduction values (from 2.8 to 5.9-log units) and was the only isolate able to inhibit E. coli O157:H7 at refrigeration temperatures on both fruits. Therefore, CPA-7 was selected for further assays. Dose-response assays showed that CPA-7 should be present in at least the same amount as the pathogen to adequately reduce the numbers of the pathogen. From the results obtained in in vitro assays, competition seemed to be CPA-7's mode of action against E. coli O157:H7. The CPA-7 strain was identified as Pseudomonas graminis. Thus, the results support the potential use of CPA-7 as a bioprotective agent against foodborne pathogens in minimally processed fruit. Copyright © 2013 Elsevier Ltd. All rights reserved.

Comparative Effect of Heat Shock on Survival of O157:H7 and Non-O157 Shiga Toxigenic Escherichia coli and Salmonella in Lean Beef with or without Moisture-Enhancing Ingredients.

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Vasan, Akhila; Ingham, Steven C; Ingham, Barbara H

2017-06-01

Thermal tolerance of pathogenic bacteria has been shown to increase after exposure to sublethal elevated temperatures, or heat shock. We evaluated the effect of heat shock at 48°C on thermal tolerance (D 55°C ) of cocktails of O157 and non-O157 Shiga toxigenic Escherichia coli (STEC) and Salmonella in lean ground beef with or without moisture-enhancing ingredients. Beef was moisture enhanced to 110% (w) with a 5% NaCl-2.5% sodium tripolyphosphate (w/w) brine. Meat, with or without added brine, was inoculated (∼10 8 CFU/g) and heat shocked at 48°C for 0, 5, or 30 min, followed by isothermal heating at 55°C. Inoculated control samples were unenhanced and were not subject to heat shock. From the linear portion of the log CFU per gram surviving cells over time plots, D 55°C -values (minutes) were calculated. D 55°C was 20.43, 28.78, and 21.15 min for O157, non-O157, and Salmonella controls, respectively. Overall, heat shock significantly increased D 55°C , regardless of pathogen (P moisture-enhanced meat (P Moisture-enhancing ingredients significantly increased D 55°C , regardless of pathogen (P moisture-enhanced beef products.

Use of Bacteriophages to Control Escherichia coli O157:H7 in Domestic Ruminants, Meat Products, and Fruits and Vegetables.

Science.gov (United States)

Wang, Lili; Qu, Kunli; Li, Xiaoyu; Cao, Zhenhui; Wang, Xitao; Li, Zhen; Song, Yaxiong; Xu, Yongping

2017-09-01

Escherichia coli O157:H7 is an important foodborne pathogen that causes severe bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Ruminant manure is a primary source of E. coli O157:H7 contaminating the environment and food sources. Therefore, effective interventions targeted at reducing the prevalence of fecal excretion of E. coli O157:H7 by cattle and sheep and the elimination of E. coli O157:H7 contamination of meat products as well as fruits and vegetables are required. Bacteriophages offer the prospect of sustainable alternative approaches against bacterial pathogens with the flexibility of being applied therapeutically or for biological control purposes. This article reviews the use of phages administered orally or rectally to ruminants and by spraying or immersion of fruits and vegetables as an antimicrobial strategy for controlling E. coli O157:H7. The few reports available demonstrate the potential of phage therapy to reduce E. coli O157:H7 carriage in cattle and sheep, and preparation of commercial phage products was recently launched into commercial markets. However, a better ecological understanding of the phage E. coli O157:H7 will improve antimicrobial effectiveness of phages for elimination of E. coli O157:H7 in vivo.

Isolation and identification of Aeromonas caviae strain KS-1 as TBTC- and lead-resistant estuarine bacteria.

Science.gov (United States)

Shamim, Kashif; Naik, Milind Mohan; Pandey, Anju; Dubey, Santosh Kumar

2013-06-01

Tributyltin chloride (TBTC)- and lead-resistant estuarine bacterium from Mandovi estuary, Goa, India was isolated and identified as Aeromonas caviae strain KS-1 based on biochemical characteristics and FAME analysis. It tolerates TBTC and lead up to 1.0 and 1.4 mM, respectively, in the minimal salt medium (MSM) supplemented with 0.4 % glucose. Scanning electron microscopy clearly revealed a unique morphological pattern in the form of long inter-connected chains of bacterial cells on exposure to 1 mM TBTC, whereas cells remained unaltered in presence of 1.4 mM Pb(NO₃)₂ but significant biosorption of lead (8 %) on the cell surface of this isolate was clearly revealed by scanning electron microscopy coupled with energy dispersive X-ray spectroscopy. SDS-PAGE analysis of whole-cell proteins of this lead-resistant isolate interestingly demonstrated three lead-induced proteins with molecular mass of 15.7, 16.9 and 32.4 kDa, respectively, when bacterial cells were grown under the stress of 1.4 mM Pb (NO₃)₂. This clearly demonstrated their possible involvement exclusively in lead resistance. A. caviae strain KS-1 also showed tolerance to several other heavy metals, viz. zinc, cadmium, copper and mercury. Therefore, we can employ this TBTC and lead-resistant bacterial isolate for lead bioremediation and also for biomonitoring TBTC from lead and TBTC contaminated environment.

Pathogenic Strains of Yersinia enterocolitica Isolated from Domestic Dogs (Canis familiaris) Belonging to Farmers Are of the Same Subtype as Pathogenic Y. enterocolitica Strains Isolated from Humans and May Be a Source of Human Infection in Jiangsu Province, China ▿ ‡

Science.gov (United States)

Wang, Xin; Cui, Zhigang; Wang, Hua; Tang, Liuying; Yang, Jinchuan; Gu, Ling; Jin, Dong; Luo, Longze; Qiu, Haiyan; Xiao, Yuchun; Xiong, Haiping; Kan, Biao; Xu, Jianguo; Jing, Huaiqi

2010-01-01

We isolated 326 Yersinia enterocolitica strains from 5,919 specimens from patients with diarrhea at outpatient clinics, livestock, poultry, wild animals, insect vectors, food, and the environment in the cities of Nantong and Xuzhou in Jiangsu Province, China, from 2004 to 2008. The results showed that the 12 pathogenic strains were of the O:3 serotype. Six strains were isolated from domestic dogs (Canis familiaris) belonging to farmers and were found to be the primary carriers of pathogenic Y. enterocolitica strains, especially in Xuzhou. Pulsed-field gel electrophoresis analysis of the pathogenic strains from dogs belonging to farmers showed that they shared the same patterns as strains from diarrhea patients isolated in 1994. This indicates that the strains from domestic dogs have a close correlation with the strains causing human infections. PMID:20181899

Collagen-like proteins in pathogenic E. coli strains.

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Neelanjana Ghosh

Full Text Available The genome sequences of enterohaemorrhagic E. coli O157:H7 strains show multiple open-reading frames with collagen-like sequences that are absent from the common laboratory strain K-12. These putative collagens are included in prophages embedded in O157:H7 genomes. These prophages carry numerous genes related to strain virulence and have been shown to be inducible and capable of disseminating virulence factors by horizontal gene transfer. We have cloned two collagen-like proteins from E. coli O157:H7 into a laboratory strain and analysed the structure and conformation of the recombinant proteins and several of their constituting domains by a variety of spectroscopic, biophysical, and electron microscopy techniques. We show that these molecules exhibit many of the characteristics of vertebrate collagens, including trimer formation and the presence of a collagen triple helical domain. They also contain a C-terminal trimerization domain, and a trimeric α-helical coiled-coil domain with an unusual amino acid sequence almost completely lacking leucine, valine or isoleucine residues. Intriguingly, these molecules show high thermal stability, with the collagen domain being more stable than those of vertebrate fibrillar collagens, which are much longer and post-translationally modified. Under the electron microscope, collagen-like proteins from E. coli O157:H7 show a dumbbell shape, with two globular domains joined by a hinged stalk. This morphology is consistent with their likely role as trimeric phage side-tail proteins that participate in the attachment of phage particles to E. coli target cells, either directly or through assembly with other phage tail proteins. Thus, collagen-like proteins in enterohaemorrhagic E. coli genomes may have a direct role in the dissemination of virulence-related genes through infection of harmless strains by induced bacteriophages.

Primary isolation strain determines both phage type and receptors recognised by Campylobacter jejuni bacteriophages.

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Martine C Holst Sørensen

Full Text Available In this study we isolated novel bacteriophages, infecting the zoonotic bacterium Campylobacter jejuni. These phages may be used in phage therapy of C. jejuni colonized poultry to prevent spreading of the bacteria to meat products causing disease in humans. Many C. jejuni phages have been isolated using NCTC12662 as the indicator strain, which may have biased the selection of phages. A large group of C. jejuni phages rely on the highly diverse capsular polysaccharide (CPS for infection and recent work identified the O-methyl phosphoramidate modification (MeOPN of CPS as a phage receptor. We therefore chose seven C. jejuni strains each expressing different CPS structures as indicator strains in a large screening for phages in samples collected from free-range poultry farms. Forty-three phages were isolated using C. jejuni NCTC12658, NCTC12662 and RM1221 as host strains and 20 distinct phages were identified based on host range analysis and genome restriction profiles. Most phages were isolated using C. jejuni strains NCTC12662 and RM1221 and interestingly phage genome size (140 kb vs. 190 kb, host range and morphological appearance correlated with the isolation strain. Thus, according to C. jejuni phage grouping, NCTC12662 and NCTC12658 selected for CP81-type phages, while RM1221 selected for CP220-type phages. Furthermore, using acapsular ∆kpsM mutants we demonstrated that phages isolated on NCTC12658 and NCTC12662 were dependent on the capsule for infection. In contrast, CP220-type phages isolated on RM1221 were unable to infect non-motile ∆motA mutants, hence requiring motility for successful infection. Hence, the primary phage isolation strain determines both phage type (CP81 or CP220 as well as receptors (CPS or flagella recognised by the isolated phages.

Misidentification of Vibrio cholerae O155 isolated from imported shrimp as O serogroup O139 due to cross-agglutination with commercial O139 antisera

DEFF Research Database (Denmark)

Dalsgaard, A.; Mazur, J.; Dalsgaard, Inger

2002-01-01

. The strain contained two plasmids, in contrast to other O139 strains, which normally do not contain plasmids. The characteristics of the strain led to further agglutination testing with other antisera that are not commercially available, and the strain was found to agglutinate O155 antiserum in repeated...... was isolated from imported raw frozen shrimp. The toxigenicity of the strain was analyzed, and the results of a polymerase chain reaction showed that the V. cholerae strain did not contain the virulence genes ctx, tcp9, and zot, which are normally found in V. cholerae O1 and O139. The strain was resistant...... to colistin and spectinomycin. The high susceptibility of the strain to antimicrobial agents was confirmed by the lack of an SXT element, a self-transmissible, chromosomal genetic element that is normally present in O139 strains and encodes resistance to sulfonamides, trimethoprim, and streptomycin...

Isolation of a potentially probiotic Lactobacillus plantarum from Siahmezgi cheese and its characterization as a potentially probiotic

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Hojjatolah Zamani

2016-03-01

β- galactosidase and hemolytic activity as well as antibiotic susceptibility. In addition, antibacterial activity of the isolated strains against E. coli O157 and Salmonella enterica serovar typhimurium ATCC 14028 was determined. Results: One strain, labeled as Lb3 showed the highest tolerance to low pH, bile and simulated gastrointestinal tract conditions. This strain exhibited resistance to Streptomycin, Vancomycin and Polymixin B as well as effective antibacterial activity against two Gram negative pathogens, lacking hemolytic activity as well as high β- galactosidase activity. Finally, the strain Lb3 was identified as Lactobacillus plantarum CJLP55 using biochemical characterization and 16S rRNA sequencing assay. Discussion and conclusion: In the present work, a potentially probiotic Lactobacillus plantarum CJLP55 was isolated from traditionally produced Siahmezgi cheese. The bacterium displayed good probiotic properties and could be used in dairy industry.

Development, validation, and standardization of polymerase chain reaction-based detection of E-coli O157

DEFF Research Database (Denmark)

Abdulmawjood, A.; Bulte, M.; Roth, S.

2004-01-01

A diagnostic polymerase chain reaction assay was developed for the detection of E. coli O157 as the first part of a multicenter validation and standardization project. The assay is based on amplification of sequences of the rfbE O157 gene and includes an internal amplification control. The select...

Comparative genomic analysis of two isolates of Vibrio cholerae O1 Ogawa El Tor isolated during outbreak in Mariupol in 2011.

Science.gov (United States)

Kuleshov, Konstantin V; Kostikova, Anna; Pisarenko, Sergey V; Kovalev, Dmitry A; Tikhonov, Sergey N; Savelievа, Irina V; Saveliev, Vilory N; Vasilieva, Oksana V; Zinich, Liliia S; Pidchenko, Nadiia N; Kulichenko, Alexander N; Shipulin, German A

2016-10-01

Cholera is a water-borne, severe enteric infection essentially caused by toxigenic strains of Vibrio cholera O1 and O139 serogroups. An outbreak of cholera was registered during May-July 2011 in Mariupol, Ukraine, with 33 cholera cases and 25 carriers of cholera. Following this outbreak, the toxigenic strain of V. cholerae 2011EL-301 was isolated from seawater in the recreation area of Taganrog city on the territory of Russia. The aim of our study was to understand genomic features of Mariupol isolates as well as to evaluate hypothesis about possible interconnection between the outbreak of cholera in Mariupol and the single case of isolation of V. cholerae from the Sea of Azov in Russia. Mariupol isolates were phenotypically characterized and subsequently subjected to whole genome sequencing procedure. Phylogenetic analysis based on high-quality SNPs of V. cholera O1 El Tor isolates of the 7th pandemic clade from different regions showed that clinical and environmental isolates from Mariupol outbreak were attributable to a unique phylogenetic clade within wave 3 of V. cholera O1 El Tor isolates and characterized by six clade-specific SNPs. Whereas Taganrog isolate belonged to distantly related clade which allows us to reject the hypothesis of transmission the outbreak strain of V. cholerae O1 from Ukraine to Russia in 2011. Mariupol isolates shared a common ancestor with Haiti\\Nepal-4\\India clade indicating that outbreak progenitor strain most likely originated in the South Asia region and later was introduced to Ukraine. Moreover, genomic data both based on hqSNPs and similarity of virulence-associated mobile genomic elements of Mariupol isolates suggests that environmental and clinical isolates are a part of joint outbreak which confirms the role of contaminated domestic sewage, as an element of the complex chain of infection spread during cholera outbreak. In general, the genome-wide comparative analysis of both genes and genomic regions of epidemiological

An aptamer-based biosensor for colorimetric detection of Escherichia coli O157:H7.

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Wenhe Wu

Full Text Available BACKGROUND: An aptamer based biosensor (aptasensor was developed and evaluated for rapid colorimetric detection of Escherichia coli (E. coli O157:H7. METHODOLOGY/PRINCIPAL FINDINGS: The aptasensor was assembled by modifying the truncated lipopolysaccharides (LPS-binding aptamer on the surface of nanoscale polydiacetylene (PDA vesicle using peptide bonding between the carboxyl group of the vesicle and the amine group of the aptamer. Molecular recognition between E. coli O157:H7 and aptamer at the interface of the vesicle lead to blue-red transition of PDA which was readily visible to the naked eyes and could be quantified by colorimetric responses (CR. Confocal laser scanning microscope (CLSM and transmission electron microscopy (TEM was used to confirm the specific interactions between the truncated aptamer and E. coli O157:H7. The aptasensor could detect cellular concentrations in a range of 10(4~ 10(8 colony-forming units (CFU/ml within 2 hours and its specificity was 100% for detection of E. coli O157:H7. Compared with the standard culture method, the correspondent rate was 98.5% for the detection of E. coli O157:H7 on 203 clinical fecal specimens with our aptasensor. CONCLUSIONS: The new aptasensor represents a significant advancement in detection capabilities based on the combination of nucleic acid aptamer with PDA vesicle, and offers a specific and convenient screening method for the detection of pathogenic bacteria. This technic could also be applied in areas from clinical analysis to biological terrorism defense, especially in low-resource settings.

[VNTR-genotyping of Vibrio cholerae strains isolated from objects in the territory of Russian Federation in 2012].

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Vodop'ianov, A S; Mazrukho, A B; Vodop'ianov, S O; Mishan'kin, B N; Kruglikov, V D; Apkhangel'skaia, I V; Oleĭnikov, I P; Zubkova, D A; Monakhova, E V; Grigorenko, L V

2014-01-01

VNTR-typing of Vibrio cholerae strains isolated in the territory of Russian Federation in 2012. 71 Vibrio cholerae O3 and 3 V cholerae O1/O139 strains were used in the study. Genotyping was performed by using PCR for 5 VNTR-loci. Multilocus VNTR-typing allowed to group the strains into 31 VNTR-genotypes. Genotypes were divided among 10 discrete clusters by results of a cluster analysis. The presence of tcpA gene is clearly linked with the presence of VcB locus. Each geographic region was characterized by their own VNTR-genotypes. In the course of the carried out VNTR-genotyping of V. cholerae isolated in 2012, 2 types of vibrio population formation were detected. A geographic attachment to specific regions was characteristic for most of the genotypes.

Assumptions of acceptance sampling and the implications for lot contamination: Escherichia coli O157 in lots of Australian manufacturing beef.

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Kiermeier, Andreas; Mellor, Glen; Barlow, Robert; Jenson, Ian

2011-04-01

The aims of this work were to determine the distribution and concentration of Escherichia coli O157 in lots of beef destined for grinding (manufacturing beef) that failed to meet Australian requirements for export, to use these data to better understand the performance of sampling plans based on the binomial distribution, and to consider alternative approaches for evaluating sampling plans. For each of five lots from which E. coli O157 had been detected, 900 samples from the external carcass surface were tested. E. coli O157 was not detected in three lots, whereas in two lots E. coli O157 was detected in 2 and 74 samples. For lots in which E. coli O157 was not detected in the present study, the E. coli O157 level was estimated to be contaminated carton, the total number of E. coli O157 cells was estimated to be 813. In the two lots in which E. coli O157 was detected, the pathogen was detected in 1 of 12 and 2 of 12 cartons. The use of acceptance sampling plans based on a binomial distribution can provide a falsely optimistic view of the value of sampling as a control measure when applied to assessment of E. coli O157 contamination in manufacturing beef. Alternative approaches to understanding sampling plans, which do not assume homogeneous contamination throughout the lot, appear more realistic. These results indicate that despite the application of stringent sampling plans, sampling and testing approaches are inefficient for controlling microbiological quality.

The Development of a Portable SPR Bioanalyzer for Sensitive Detection of Escherichia coli O157:H7

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Shun Wang

2016-11-01

Full Text Available The purpose of this study was to develop a portable surface plasmon resonance (SPR bioanalyzer for the sensitive detection of Escherichia coli O157:H7 in comparison with an enzyme-linked immunosorbent assay (ELISA. The experimental setup mainly consisted of an integrated biosensor and a homemade microfluidic cell with a three-way solenoid valve. In order to detect Escherichia coli O157:H7 using the SPR immunoassay, 3-mercaptopropionic acid (3-MPA was chemisorbed onto a gold surface via covalent bond for the immobilization of biological species. 1-ethyl-3-(3-dimethylaminopropyl carbodiimide hydrochloride (EDC and N-hydroxysuccinimide (NHS were used as crosslinker reagents to enable the reaction between 3-MPA and Escherichia coli O157:H7 antibodies by covalent –CO–NH– amide bonding. The experimental results were obtained from the Escherichia coli O157:H7 positive samples prepared by 10-, 20-, 40-, 80-, and 160-fold dilution respectively, which show that a good linear relationship with the correlation coefficient R of 0.982 existed between the response units from the portable SPR bioanalyzer and the concentration of Escherichia coli O157:H7 positive samples. Moreover, the theoretical detection limit of 1.87 × 103 cfu/mL was calculated from the positive control samples. Compared with the Escherichia coli O157:H7 ELISA kit, the sensitivity of this portable SPR bioanalyzer is four orders of magnitude higher than the ELISA kit. The results demonstrate that the portable SPR bioanalyzer could provide an alternative method for the quantitative and sensitive determination of Escherichia coli O157:H7 in field.

Antibacterial activities of wasabi against Escherichia coli O157:H7 and Staphylococcus aureus

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Zhongjing Lu

2016-09-01

Full Text Available Escherichia coli O157:H7 and Staphylococcus aureus are two of the major pathogens frequently involved in foodborne outbreaks. Control of these pathogens in foods is essential to food safety. It is of great interest in the use of natural antimicrobial compounds present in edible plants to control foodborne pathogens as consumers prefer more natural green foods. Allyl isothiocyanate (AITC is an antimicrobial compound naturally present in wasabi (Japanese horseradish and several other edible plants. Although the antibacterial effects of pure AITC and wasabi extract (essential oil against several bacteria have been reported, the antibacterial property of natural wasabi has not been well studied. This study investigated the antibacterial activities of wasabi as well as AITC against E. coli O157:H7 and S. aureus. Chemical analysis showed that AITC is the major isothiocyanate in wasabi. The AITC concentration in the wasabi powder used in this study was 5.91±0.59 mg/g. The minimum inhibitory concentration (MIC of wasabi against E. coli O157:H7 or S. aureus was 1% (or 10 mg/ml. Wasabi at 4% displayed higher bactericidal activity against S. aureus than against E. coli O157:H7. The MIC of AITC against either pathogen was between 10 and 100 µg/ml. AITC at 500 µg/ml was bactericidal against both pathogens while AITC at 1000 µg/ml eliminated E. coli O157:H7 much faster than S. aureus. The results from this study showed that wasabi has strong antibacterial property and has high potential to effectively control E. coli O157:H7 and S. aureus in foods. The antibacterial property along with its natural green color, unique flavor, and advantage to safeguard foods at the point of ingestion makes wasabi a promising natural edible antibacterial plant. The results from this study may be of significant interest to the food industry as they develop new and safe foods. These results may also stimulate more research to evaluate the antibacterial effect of wasabi

Antibacterial Activities of Wasabi against Escherichia coli O157:H7 and Staphylococcus aureus.

Science.gov (United States)

Lu, Zhongjing; Dockery, Christopher R; Crosby, Michael; Chavarria, Katherine; Patterson, Brett; Giedd, Matthew

2016-01-01

Escherichia coli O157:H7 and Staphylococcus aureus are two of the major pathogens frequently involved in foodborne outbreaks. Control of these pathogens in foods is essential to food safety. It is of great interest in the use of natural antimicrobial compounds present in edible plants to control foodborne pathogens as consumers prefer more natural "green" foods. Allyl isothiocyanate (AITC) is an antimicrobial compound naturally present in wasabi (Japanese horseradish) and several other edible plants. Although the antibacterial effects of pure AITC and wasabi extract (essential oil) against several bacteria have been reported, the antibacterial property of natural wasabi has not been well studied. This study investigated the antibacterial activities of wasabi as well as AITC against E . coli O157:H7 and S . aureus . Chemical analysis showed that AITC is the major isothiocyanate in wasabi. The AITC concentration in the wasabi powder used in this study was 5.91 ± 0.59 mg/g. The minimum inhibitory concentration (MIC) of wasabi against E. coli O157:H7 or S. aureus was 1% (or 10 mg/ml). Wasabi at 4% displayed higher bactericidal activity against S. aureus than against E. coli O157:H7. The MIC of AITC against either pathogen was between 10 and 100 μg/ml. AITC at 500 μg/ml was bactericidal against both pathogens while AITC at 1000 μg/ml eliminated E. coli O157:H7 much faster than S. aureus . The results from this study showed that wasabi has strong antibacterial property and has high potential to effectively control E. coli O157:H7 and S. aureus in foods. The antibacterial property along with its natural green color, unique flavor, and advantage to safeguard foods at the point of ingestion makes wasabi a promising natural edible antibacterial plant. The results from this study may be of significant interest to the food industry as they develop new and safe foods. These results may also stimulate more research to evaluate the antibacterial effect of wasabi against other

A rapid two dot filter assay for the detection of E. coli O157 in water samples.

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Kamma, Sujatha; Tang, Lily; Leung, Kelvin; Ashton, Edie; Newman, Norman; Suresh, Mavanur R

2008-07-31

E. coli O157:H7 is an enterohemorrhagic bacteria that cause deadly water-borne infections implicated in outbreaks of a wide spectrum of human gastrointestinal diseases. It is therefore important to have a rapid convenient, simple and sensitive range of detection of E. coli O157:H7. A new E. coli O157 MAb designated P124 was developed for ultrasensitive detection of E. coli O157 in water, apple juice and beef for routine use. A prototype filter dot assay was designed with anti-E. coli O157 MAb bound to 0.2 microm nitrocellulose filter disk as the capture antibody. A 100 ml water sample spiked with 1-50 CFU of E. coli O157 either in the presence or absence of other non-specific bacteria were filtered for capture of the pathogen on the antibody coated nitrocellulose disk. The detection of the pathogen was successfully accomplished by the same antibody both as a capture and detecting antibody as a homosandwich. In a non-enriched format, detection of E. coli was possible with a sensitivity of 2500 CFU/100 ml. Ultrasensitive detection of ~1 CFU/100 ml sample could be achieved by a prior pathogen enrichment step before the addition of the labeled antibody. The design of this diagnostic test is based on the common architecture of all bacteria, viruses and spores, namely the manifestation of repeat lipopolysaccharide epitopes on the surface. We have developed an easy-to-use two dot visual filter assay for translation into current water testing in public health laboratories to detect E. coli O157:H7. In a 5 h assay approximately 1 CFU and approximately 5 CFU of E. coli O157 could be detected in 100 ml of water or juice and lake samples respectively. This simple homosandwich enrichment strategy can also be used to detect low levels of other water-borne pathogens.

Water sources as reservoirs of Vibrio cholerae O1 and non-O1 strains in Bepanda, Douala (Cameroon): relationship between isolation and physico-chemical factors.

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Akoachere, Jane-Francis Tatah Kihla; Mbuntcha, Christelle Kwedjeu Pulcherie

2014-07-30

Cholera has been endemic in Douala since 1971. Most outbreaks start from Bepanda, an overcrowded neighbourhood with poor hygiene and sanitary conditions. We investigated water sources in Bepanda as reservoirs of Vibrio cholerae, the causative agent of cholera, determined its antibiotic susceptibility and some physico-chemical characteristics that could maintain the endemicity of this organism in Bepanda. Three hundred and eighteen water samples collected from 45 wells, 8 taps and 1 stream from February to July 2009 were analyzed for V. cholerae using standard methods. Isolates were characterized morphologically, biochemically and serologically. The disc diffusion technique was employed to investigate antibiotic susceptibility. Differences in prevalence of organism between seasons were analysed. Correlation strength and direction of association between physico-chemical parameters and occurrence of V. cholerae was analyzed using the Kendall tau_b non-parametric correlation. This was further confirmed with the forward-stepwise binary logistic regression. Eighty-seven (27.4%) samples were positive for V. cholerae. Isolation was highest from wells. The organism was isolated in the rainy season and dry season but the frequency of isolation was significantly higher (χ2 = 7.009, df = 1, P = 0.008) in the rainy season. Of the 96 confirmed V. cholerae isolates, 32 (33.3%) belonged to serogroup O1 and 64 (66.6%) were serogroup non-O1/non-O139. Isolates from tap (municipal water) were non-O1/non-O139 strains. Salinity had a significant positive correlation with isolation in the dry season (+0.267, P = 0.015) and rainy season (+0.223, P = 0.028). The forward-stepwise method of binary logistic regression indicated that as pH (Wald = 11.753, df = 1), P = 0.001) increased, odds of isolation of V. cholerae also increased (B = 1.297, S.E = 0.378, Exp(B) = 3.657). All isolates were sensitive to ciprofloxacin and ofloxacin. Multi-drug resistance was predominant among the non-O1/non-O