Susceptibility of Escherichia coli O157 and non-O157 isolates to lactate

AIMS: To examine the effect of temperature on the antimicrobial efficacy of lactate and propionate against O157 and non-O157 Escherichia coli isolates. METHODS AND RESULTS: Lactate and, to a lesser extent, propionate effectively reduced viability at 37 degrees C. Ethanol enhanced this effect. Reducing the temperature to 20 or 5 degrees C caused an increase in survival in the presence of these organic acids with or without ethanol. At 20 degrees C the deltapH, membrane potential and intracellular lactate anion concentration were less than at 37 degrees C. CONCLUSIONS: The efficacy of lactate and propionate against E. coli O157 and non-O157 isolates is reduced at lower temperatures, perhaps due to the reduction in the deltapH, membrane potential and intracellular lactate anion concentration. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings suggest that the usefulness of organic acids as decontaminants for E. coli O157 is temperature dependent.     

Methods available for the detection of Escherichia coli O157 in clinical,food and environmental samples

Because of the potential severity of infections caused by Escherichia coli O157 it is important that the most sensitive laboratory methods are used both for outbreak investigation and surveillance. Selective culture of E. coli O157 remains the detection method of choice, particularly in investigation of outbreaks where strains isolated from various sources may need to be compared by various typing methods. Strains of E. coli O157 do not normally ferment sorbitol, whereas many other serogroups of E. coli do, and sorbitol MacConkey agar, or modified forms of this medium, have become widely used for their isolation. Detection of small numbers of E. coli O157 may be facilitated by enrichment culture which may include a recovery period during which selective agents are not added to the medium. Immunomagnetic separation of E. coli O157 after enrichment culture enhances sensitivity still further and has the potential to be fully automated. Alternatives to culture include immunoassays and PCR, both of which are available as commercial detection kits. The last 15 years has seen many advances in detection of E. coli O157 and has been accompanied by a plethora of reports in the scientific literature. However, it is an area which is continually developing and we are still far away from a universally accepted method for this purpose. This revised version was published online in November 2006 with corrections to the Cover Date.     

Waterborne Escherichia coli O157

The waterborne route of Vero cytotoxin-producing E. coli (VTEC) O157 infection was first suggested in two unconnected human cases in 1985. Since then, waterborne VTEC O157 has been identified in sporadic cases and in outbreaks of illness. Recreational waters, private and municipal supplies have been implicated from microbiological, environmental and epidemiological studies of cases. In addition, a research cohort study of farm workers identified exposure to private water supplies as a risk factor for having antibodies to E. coli O157. Sources of contamination are thought to be animal and human faeces or sewage. The presence of low numbers of target organisms in water makes microbiological confirmation difficult, therefore epidemiological evidence has been essential in outbreak investigations. Despite the potential for contamination of water with VTEC O157, waterborne infection is relatively rare largely due to the susceptibility of the organism to water treatment processes. This paper presents the evidence for waterborne VTEC O157 infection, considering current microbiological, environmental and particularly epidemiological information.     

Escherichia coli O157 and non-O157 isolates are more susceptible to L-lactate than to D-lactate

The antimicrobial effect of L-lactate was much greater than that of D-lactate over a range of concentrations for Escherichia coli O157 and non-O157 strains. Despite this, the intracellular pHs and membrane potentials of L-lactate- and D-lactate-treated cells were similar, suggesting that these factors are not involved in the antimicrobial action of L-lactate.     

Comparison of traditional and molecular typing methods of Escherichia coli O157

An account is given using typing methods and detection of virulence genes of different serotypes of Escherichia coli isolated in Hungary. By hybridization using SLT-I and SLT-II probes and PCR method using stx1-2, eae and ehx primers we could differentiate O157 strains of different serotypes into eight (stx, eae, ehxA positive; stx, eae positive; stx, ehxA positive; stx positive; eae, ehxA positive; eae positive; ehxA positive; stx, eae, ehxA negative) types. The discriminatory power of phage typing proves to be much higher than that of the plasmid profile. RAPD typing with different primers could confirm or exclude the subtypes identity of the isolated E. coli O157 serotypes. Escherichia coli O157:HNM isolates could be sorted in six different phage types and six different RAPD types with ERIC-1, in five RAPD types with ERIC-2 and in seven types with M13 primers. Escherichia coli O157:H7 showed six different phage types and three RAPD types with ERIC-1 and ERIC-2 and five types with M13 primers. According to our results the standard PFGE protocol [32] gives the opportunity to differentiate epidemiologically independent but evolutionary related or unrelated isolates, but the practical value of PFGE method for epidemiological purposes must be confirmed by other or more restriction enzymes or using an other protocol. Summarizing our results we suggest the use of phage and RAPD typing and in doubtful cases the PFGE method.     

Release of membrane vesicles containing endotoxic lipopolysaccharide in Escherichia coli O157:H7 clinical isolates

Membrane vesicles released by E. coli O157:H7 strains were investigated by immuno-electron microscopy using anti-O157 antibody. Anti-O157 antibody enhanced the negative-staining of vesicles and we found numerous small vesicles clearly formed around bacterial cells. An immunogold-electron microscopic examination confirmed that lipopolysaccharide (LPS) including the O-side chain is present on the surface of the vesicles. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of the purified vesicles showed that the vesicles contained LPS consisting of a lipid-A and an O polysaccharide. In addition, the endotoxic activity of the vesicle was confirmed by a limulus test. These results suggest that the vesicles may play an important role in the pathogenesis of Escherichia coli O157:H7.     

Biocontrol of Escherichia coli O157 with O157-Specific Bacteriophages

Escherichia coli O157 antigen-specific bacteriophages were isolated and tested to determine their ability to lyse laboratory cultures of Escherichia coli O157:H7. A total of 53 bovine or ovine fecal samples were enriched for phage, and 5 of these samples were found to contain lytic phages that grow on E. coli O157:H7. Three bacteriophages, designated KH1, KH4, and KH5, were evaluated. At 37 or 4°C, a mixture of these three O157-specific phages lysed all of the E. coli O157 cultures tested and none of the non-O157 E. coli or non-E. coli cultures tested. These results required culture aeration and a high multiplicity of infection. Without aeration, complete lysis of the bacterial cells occurred only after 5 days of incubation and only at 4°C. Phage infection and plaque formation were influenced by the nature of the host cell O157 lipopolysaccharide (LPS). Strains that did not express the O157 antigen or expressed a truncated LPS were not susceptible to plaque formation or lysis by phage. In addition, strains that expressed abundant mid-range-molecular-weight LPS did not support plaque formation but were lysed in liquid culture. Virulent O157 antigen-specific phages could play a role in biocontrol of E. coli O157:H7 in animals and fresh foods without compromising the viability of other normal flora or food quality.     

Antimicrobial resistance of Escherichia coli O157 isolated from humans, cattle, swine, and food

A total of 361 Escherichia coli O157 isolates, recovered from humans, cattle, swine, and food during the years 1985 to 2000, were examined to better understand the prevalence of antimicrobial resistance among these organisms. Based on broth microdilution results, 220 (61%) of the isolates were susceptible to all 13 antimicrobials tested. Ninety-nine (27%) of the isolates, however, were resistant to tetracycline, 93 (26%) were resistant to sulfamethoxazole, 61 (17%) were resistant to cephalothin, and 48 (13%) were resistant to ampicillin. Highest frequencies of resistance occurred among swine isolates (n = 70), where 52 (74%) were resistant to sulfamethoxazole, 50 (71%) were resistant to tetracycline, 38 (54%) were resistant to cephalothin, and 17 (24%) were resistant to ampicillin. Based on the presence of Shiga toxin genes as determined by PCR, 210 (58%) of the isolates were identified as Shiga toxin-producing E. coli (STEC). Among these, resistance was generally low, yet 21 (10%) were resistant to sulfamethoxazole and 19 (9%) were resistant to tetracycline. Based on latex agglutination, 189 (52%) of the isolates were identified as E. coli O157:H7, among which 19 (10%) were resistant to sulfamethoxazole and 16 (8%) were resistant to tetracycline. The data suggest that selection pressure imposed by the use of tetracycline derivatives, sulfa drugs, cephalosporins, and penicillins, whether therapeutically in human and veterinary medicine or as prophylaxis in the animal production environment, is a key driving force in the selection of antimicrobial resistance in STEC and non-STEC O157.     

Isolation and characteristics of sorbitol-fermenting Escherichia coli O157 strains from cattle

Cattle can be a reservoir of sorbitol-fermenting Escherichia coli O157 (SF E. coli O157) and a source of human diseases. In this study, six strains of SF E. coli O157 were isolated and characterized from cattle using an immunomagnetic separation procedure. PCR analysis of the SF E. coli O157 virulence markers showed that all six isolates tested positive for sfpA, rfbE and eaeA, and negative for terA, ureA, katP and espP. Two of the isolates contained the stx genes. Four isolates tested positive for enterohemorrhagic E. coli hlyA (EhlyA) by PCR but were nonhemolytic on the blood agar. Five isolates tested positive for the cdtA gene. The possession of these virulence factors was an indication of their pathogenic potential. The random amplified polymorphic DNA patterns, which were generated by the arbitrarily primed PCR of the SF E. coli O157 isolates from the cattle, were significantly different from those of the non-sorbitol-fermenting E. coli O157 (NSF E. coli O157) strains originating from cattle or humans. GelCompar analysis showed that the SF E. coli O157 isolates had only a 57% genetic similarity with the NSF E. coli strains. The minimal inhibitory concentration assay showed that imipenem inhibited the growth of the six isolates at a concentration of <4 microg/ml.     

Biocontrol of Escherichia coli O157 with O157-specific bacteriophages.

Escherichia coli O157 antigen-specific bacteriophages were isolated and tested to determine their ability to lyse laboratory cultures of Escherichia coli O157:H7. A total of 53 bovine or ovine fecal samples were enriched for phage, and 5 of these samples were found to contain lytic phages that grow on E. coli O157:H7. Three bacteriophages, designated KH1, KH4, and KH5, were evaluated. At 37 or 4 degrees C, a mixture of these three O157-specific phages lysed all of the E. coli O157 cultures tested and none of the non-O157 E. coli or non-E. coli cultures tested. These results required culture aeration and a high multiplicity of infection. Without aeration, complete lysis of the bacterial cells occurred only after 5 days of incubation and only at 4 degrees C. Phage infection and plaque formation were influenced by the nature of the host cell O157 lipopolysaccharide (LPS). Strains that did not express the O157 antigen or expressed a truncated LPS were not susceptible to plaque formation or lysis by phage. In addition, strains that expressed abundant mid-range-molecular-weight LPS did not support plaque formation but were lysed in liquid culture. Virulent O157 antigen-specific phages could play a role in biocontrol of E. coli O157:H7 in animals and fresh foods without compromising the viability of other normal flora or food quality.     

Heterogeneity of phenotypic and genotypic traits including organic-acid resistance in Escherichia coli O157 isolates

We undertook an epidemiologic study for the sensitivity of both Shiga-like toxin (Slt)-producing Escherichia coli (STEC) O157 and non-STEC O157 strains isolated from different patients with diarrhea to hydrochloric acid (HCl) and organic acids such as acetate, propionate, butyrate and lactate, and other pathogenic factors. The E. coli O157 isolates examined showed a wide variety of organic-acid susceptibility patterns. E. coli O157 isolates resistant to HCl or acetate were found more frequently than those resistant to other organic acids. These isolates also showed diverse pathogenicity patterns for the presence of the virulence genes, antibiotic susceptibility and plasmid profile.     

Phenotypic and molecular characteristics of Escherichia coli isolated from aquatic environment of Bangladesh

Pathogenic Escherichia coli remains important etiological agent of infantile diarrhea in Bangladesh. Previous studies have focused mostly on clinical strains, but very little is known about their presence in aquatic environments. The present study was designed to characterize potentially pathogenic E. coli isolated between November 2001 and December 2003 from aquatic environments of 13 districts of Bangladesh. Serotyping of 96 randomly selected strains revealed O161 to be the predominant serotype (19%), followed by O55 and O44 (12% each), and 11% untypable. Serotype-based pathotyping of the E. coli strains revealed 47%, 30%, and 6% to belong to EPEC, ETEC, and EHEC pathotypes, respectively. The majority of the 160 strains tested were resistant to commonly used antimicrobial agents. Plasmid pro-filing showed a total of 17 different bands ranging from 1.3 to 40 kb. However, 35% of the strains did not contain any detectable plasmid, implying no correlation between plasmid and drug resistance. Although virulence gene profiling revealed 97 (61%) of the strains to harbor the gene encoding heat-stable enterotoxin (ST), 2 for the gene encoding Shiga toxin (Stx), and none for the gene for heat-labile enterotoxin (LT), serotype-based pathotyping of E. coli was not fully supported by this gene profiling. A dendrogram derived from the PFGE patterns of 22 strains of three predominant serogroups indicated two major clusters, one containing mainly serogroup O55 and the other O8. Three strains of identical PFGE profiles belonging to serogroup O55 were isolated from three distinct areas, which may be of epidemiological significance. Finally, it may be concluded that serotype-based pathotyping may be useful for E. coli strains of clinical origin; however, it is not precise enough for reliably identifying environmental strains as diarrheagenic.     

玉溪市大肠埃希菌O157菌株的流行率、表型与毒力因子特征

目的:检测并分析大肠埃希菌O157(Escherichia coli O157)在玉溪市奶牛、猪、鸡和腹泻病人中的带菌率及其特征,为E.coli O157感染症的防治和该菌分离鉴定技术的建立提供科学依据.方法:将玉溪市最大的一个奶牛场、1个猪屠宰场、2个生禽市场和市医院检验科作为监测点,采集牛、猪、鸡、腹泻病人粪便标本数分别为70、250、350和400例,用免疫磁性分离法和免疫色层技术进行E.coli O157菌株的分离培养,纯化菌株经3种鉴别培养基培养以及法国生物梅里埃公司全自动微生物鉴定系统VITEK 32 GNI 或肠杆菌科鉴定系统API 20 E试条、血清学、噬菌体型、亚碲酸盐抗性、聚合酶链反应等检测与分析.结果:牛、猪、鸡、腹泻病人E.coli O157的带菌率分别为1.4%、2.0%、0和1.2%,分离到4株E.coli O157∶H7、6株E.coli O157∶Hund和1株E.coli O157∶HNM,认识了11株E.coli O157的形态学、生物化学、血清学、噬菌体型、亚碲酸盐抗性、毒力因子等特征.结论:单一动物群或人群可同时携带多株E.coli O157,甚至同一个体也会携带2株E.coli O157;E.coli O157菌株间的表型与毒力因子特征存在一定差异,基因指纹技术可确认并证实菌株间的差异性.     

Veterinary significance of verocytotoxin-producing Escherichia coli O157

Verocytotoxin-producing Escherichia coli O157 is a serious pathogen in man that is carried by ruminants and has been isolated from some other animal species. Except in the very young of certain species and in greyhounds, the organism is not associated with disease in animals. Humans may be infected by ingestion of the organism through direct animal contact, from contaminated food or water or from the environment. Great efforts have been made to improve hygienic food production and handling, to protect water supplies and to give adequate advice to people handling animals. It is also essential to try to reduce the numbers of organisms shed by animals and, to do this, a clear understanding of the ecology of the organism is required. This revised version was published online in November 2006 with corrections to the Cover Date.     

Clinical significance of Verocytotoxin-producing Escherichia coli O157

In 1977, Konowalchuk and colleagues (Konowalchuk, J., Speirs, J.I. & Stavric, S. 1977 Infection and Immunity 18, 775–779) were the first to describe Verocytotoxin-producing strains of Escherichia coli or VTEC. The surveillance of infection caused by VTEC demonstrated strains of E. coli belonging to serogroup O157 as the main cause of human infection capable of causing haemorrhagic colitis (HC) and haemolytic uraemic syndrome (HUS). Infection with O157 VTEC results in a range of disease manifestations including abdominal cramps, vomiting and fever. This frequently leads to cases with bloody diarrhoea and HC, and approximately 10% of patients develop HUS. The symptoms of disease caused by VTEC O157 have been well documented and the pathogenic mechanisms expressed by VTEC have been the focus of considerable attention. However, the role of putative pathogenic mechanisms in the pathogenesis of disease is not fully understood. The aim of this review is to consider the clinical aspects of infection with strains of VT-producing E. coli O157 in terms of the putative pathogenic mechanisms expressed by these bacteria. This revised version was published online in November 2006 with corrections to the Cover Date.     

Prevalence and molecular characteristics of sequence type 131 clone among clinical uropathogenic Escherichia coli isolates in Riyadh,Saudi Arabia

BackgroundThe antimicrobial resistance of extraintestinal pathogenic Escherichia coli (ExPEC) has progressively been reported worldwide. This resistance has been ascribed to global dissemination of a single E. coli clone, namely E. coli sequence type 131 (E. coli ST131). The main goal of this study is to determine the prevalence and molecular traits of ST131 and its subclones among E. coli clinical urine isolates in Riyadh, Saudi Arabia.MethodsSixty E. coli urine isolates, of different extended spectrum β-lactamase (ESBL) carriage, were involved in this study. Molecular characterization was carried out to determine the ST131 status, phylogenetic groups and virulence carriage of these isolates. ST131 isolates were further tested to evaluate the prevalence of different phylogenetic groups, subclones and virulence carriage.ResultsGroup B2 was the most common phylogroup from which E. coli isolates derived. Overall, 37 of 60 (61.7%) isolates belonged to ST131 clones. Of these, 19 (31.7%) isolates were from the H30 subclone, including 10 (16.7%) H30 non-Rx and 9 (15%) H30Rx. The remaining 18 (30%) ST131 isolates belonged to other non H30 subclones. H30 subclone was significantly higher in the virulence carriage in comparison to non H30 ST131 subclones.ConclusionThis study reported the prevalence and traits of clinical E. coli ST131 main subclones in Saudi Arabia. It also demonstrated the high prevalence of E. coli ST131 locally, and found different virulence genotypes and antimicrobial resistance phenotypes among ST131 subclones. In the future, preforming whole genome sequence-based studies on ST131 and its subclones is crucial to elucidate factors that drive the success of these organisms.     

Immunomagnetic separation of Escherichia coli O26, O111 and O157 from vegetables

AIMS: Raw fruits and vegetables have been increasingly associated with human infections caused by Shiga toxin-producing Escherichia coli. This study evaluates the isolation and detection of E. coli O26, O111 and O157 from vegetable samples using immunomagnetic particles. METHODS AND RESULTS: Standard cultivation and immunomagnetic separation (IMS) procedures were compared. It was found that immunomagnetic particles could efficiently concentrate E. coli cells, detecting significantly more bacteria than with standard cultivation procedures. CONCLUSION: Bacteria were detected in 93-100% of the inoculated samples using the IMS procedure, but only 36-93% samples tested by standard cultivation procedures were found to be positive. SIGNIFICANCE AND IMPACT OF THE STUDY: The results indicate that E. coli O26, O111 and O157 immunomagnetic particles can be a very useful and efficient tool for the detection of E. coli strains in raw vegetables, and could probably be used with samples of animal origin.     

Detection of Escherichia coli O157 in French food samples using an immunomagnetic separation method and the VIDASTME. coli O157

C. VERNOZY-ROZAND, C. MAZUY, S. RAY-GUENIOT, S. BOUTRAND-LOEï, A. MEYRAND AND y. richard. 1997. Two commercially available screening methods, an automated enzyme-linked fluorescent immunoassay (VIDAS^TM E. coli O157) and an immunomagnetic separation followed by culture onto cefixime tellurite sorbitol MacConkey agar (CT-SMAC), were compared for detection of Escherichia coli O157 in naturally and artificially contaminated food samples. A total of 250 naturally contaminated food samples, including raw milk cheeses, poultry, raw sausages and ground beef retail samples, were examined. Four poultry, one raw sausage and one ground beef sample were found to be positive for E. coli O157 by both methods. Of the six positive samples, five were shown to contain sorbitol-positive, O157-positive, H7-negative, motile and non-verotoxin-producing E. coli .