Virulence properties of Escherichia coli strains belonging to serogroups O26, O55, O111 and O128 isolated in the United Kingdom in 1991 from patients with diarrhoea.

Some strains of Escherichia coli belonging to serogroups O26, O55, O111 or O128 produce Vero cytotoxin (VT). These serogroups are included in the range of enteropathogenic E. coli (EPEC) serogroups for which commercial antisera are available. In an attempt to obtain information on VT-producing strains other than those of serogroup O157, 122 strains belonging to these four serogroups and isolated in 1991 from patients with diarrhoea in the United Kingdom were tested for hybridization with VT probes. Only 18 of the 122 strains were VT-positive and these were O26 or O128. However 90 strains hybridized with the E. coli attaching and effacing (eae) probe (including 14 VT-positive strains) and 17 with the enteroaggregative E. coli (EAggEC) probe. For 78 eae-positive and 9 EAggEC-positive strains, tissue culture tests correlated with the probe results as the strains gave, respectively, either localized adhesion and a positive fluorescent-actin staining test or a characteristic aggregative attachment. A total of 111 of the 122 strains belonging to serogroups O26, O55, O111 or O128 possessed properties that may be associated with the ability to cause human diarrhoeal disease, and similar studies are needed on strains from the other classical EPEC serogroups.     

Molecular epidemiology of Escherichia coli isolated from young South African children with diarrhoeal diseases

Molecular techniques were used for studying the epidemiology of diarrhoeal infections due to Escherichia coli in the Gauteng region in South Africa. In total, 151 E. coli strains isolated from stools of patients with diarrhoea and 30 strains isolated from stools of healthy individuals were collected between March 1996 and May 1997. The E. coli isolates were characterized by serotyping, antimicrobial susceptibility testing, and adherence patterns. Polymerase chain reaction (PCR) was performed to determine the presence of the genes-encoding virulence factors. PCR showed that 59 (32.6%) of the E. coli isolates carried eaeA genes, 6 (3.3%) possessed bfpA genes, 4 (2.2%) CNF1, and 2 (1.1%) carried labile toxin and Stx2 genes. The eae genes were more prevalent in strains isolated from patients than in those from the control group (p < 0.001). Forty-eight (26.5%) strains belonged to enteropathogenic E. coli (EPEC) O serogroups and 14 (7.7%) to Shiga toxin-producing E. coli (STEC) O157 serotype. A high percentage (28.2%) of atypical EPEC strains possessing the eaeA but not the bfpA genes was isolated. Most isolates were susceptible to commonly-used antimicrobial agents. The adherence of the E. coli strains to HeLa cells was identified more in patients (69.4%) than in the control group (60%) and was more dominant in infants than in adults. PCR and tissue culture assays were shown to be useful techniques for the epidemiological study of E. coli where this organism is a major cause of diarrhoea.     

Attaching and effacing Escherichia coli and Shiga toxin-producing E. coli in children with acute diarrhoea and controls in Teresina/PI, Brazil

This 3.5-year prospective study was conducted to ascertain the level of attaching and effacing Escherichia coli (AEEC) associated diarrhoea in children from Teresina, a northeastern state of Brazil. Passed faecal specimens from 400 patients (250 with and 150 without diarrhoea) up to 60 months of age attending from 2004 to 2007 at two public hospitals were investigated. Conventional microbiology methods and PCR were employed. Escherichia coli was isolated from 390 children, 240 of them with diarrhoea. A total of 117 AEEC strains were cultivated from specimens from 63 children, 37 with and 26 without diarrhoea. No association between AEEC and diarrhoea was observed. Atypical enteropathogenic E. coli (a-EPEC) (79.4%) was more commonly found than typical EPEC (t-EPEC). Association between EPEC and EPEC subtypes and diarrhoea was not detected. Mixed infection by t-EPEC and a-EPEC and infection by Shiga toxin-producing E. coli (STEC) were rare. Enteropathogenic E. coli was more common in males and in children aged less than 12 months. Correlation between serotyping and PCR results was 0.19. High resistance rates of AEEC to ampicillin, cephalotin, and trimethoprim-sulfamethoxazole were found. In conclusion, EPEC is very common in children with diarrhoea and controls in the population we studied, with a-EPEC predominating. This diarrhoeagenic E. coli (DEC) pathotype is more common in infant males and is resistant to drugs frequently used in clinical practice.     

Rapid presumptive identification of enteropathogenic Escherichia coli in faecal smears by means of fluorescent antibody: 3. Field evaluation

This paper reports on the evaluation of the fluorescent antibody technique for detection of enteropathogenic Escherichia coli in faecal smears from children both with and without diarrhoea. In field studies involving 315 children in Puerto Rico, it was demonstrated that presumptive diagnosis of infection with enteropathogenic E. coli could be made more rapidly and with greater sensitivity by immunofluorescence than by isolation and slide agglutination. The specificity of the fluorescent antibody test was of the same order as that of slide agglutination tests with OB antisera. The incidence of salmonellae, shigellae, coagulasepositive staphylococci and Candida in the diarrhoeal specimens was studied, but no relationship appeared to exist between these organisms and the enteropathogenic E. coli.     

The association of enterotoxigenic and enteropathogenic Escherichia coli and other enteric pathogens with childhood diarrhoea in Yugoslavia

The presence of enterotoxigenic and enteropathogenic Escherichia coli (ETEC and EPEC, respectively) was investigated in stool specimens of 1082 preschool children with diarrhoea and in stools of 335 healthy controls in localities in southern Yugoslavia, as well as in 566 children with diarrhoea and in 231 controls living in northern part of the country, during the seasonal peak (August-November) of enteric diseases in 1986. ETEC were found in 114 (10.5%) children with diarrhoea and in 14 (4.2%) controls (P less than 0.001) in the southern part, and in 26 (4.6%) ill children and one (0.4%) well child (P less than 0.005) in the northern part of Yugoslavia. EPEC were isolated from stools of 85 (7.9%) children with diarrhoea and of 14 (4.2%) well children (P less than 0.05) in localities of southern Yugoslavia, and from 22 (3.9%) ill children and from 10 (4.3%) controls in northern Yugoslavia. Nineteen EPEC strains expressed localized adherence to HEp-2 tissue culture cells; all were isolated from stools of ill children. In southern Yugoslavia, where other enteropathogens were sought, the most commonly found agents in ill children were shigellae (17.5%), rotavirus (11.8%), ETEC, and EPEC. Potential pathogens were detected in 44.5% cases of sporadic diarrhoea and in 15.8% controls. This study revealed that ETEC were associated with acute diarrhoeal disease in Yugoslav preschool children. On the other hand, the diagnosis of EPEC-diarrhoea by routine determination of serogroup established the association of these agents with sporadic diarrhoea only in the 0-2 years age categories in all investigated localities. In the less developed southern part of Yugoslavia bacteria were the predominant causative agents of enteric illness during the seasonal peak of this disease.     

病原性大肠埃希菌腹泻的实验与临床研究

目的：探索病原性大肠埃希菌腹泻的实验与临床关系。方法：采集临床门诊急性腹泻患者粪便，进行ETEC，EPEC，EIEC等病原菌常规检测，并收集患者病史临床表现。结果：从839例腹泻患者中检出病原性大肠埃希菌235株，检出率为28．01％；其中ETEC 118株（14．06％），。EPEC 93株（11．08％），EIEC 24株（2．86％），占病原菌检出的61．84％。对214印例病原性大肠埃希菌腹泻患者病史临床表现分析，3种病原性大肠埃希菌引起腹泻，临床表现不尽相同。病原性大肠埃希菌问或同其它病原菌混合感染43例13．5％），其临床表现，同单一病原菌感染比较，差异无显著性。结论：病原性大肠埃希菌引起腹泻的临床表现差异甚大；实验室病原检测是重要依据；对腹泻病因诊断有重要参考意义。     

Isolation and serological identification of enteropathogenic Escherichia coli in pasteurized milk in Brazil

OBJECTIVE: To evaluate the microbiological quality of pasteurized milk commercialized in Rio de Janeiro, Brazil, and determine serologically enteropathogenic Escherichia coli (EPEC) strains in E. coli isolates obtained from milk samples. METHODS: Ninety samples of pasteurized milk - types B and C - of three different commercial brands, purchased in supermarkets and bakeries in Rio de Janeiro, were examined. The amount of total and fecal coliform bacteria was estimated using the Most Probable Number technique. Mesophilic, psychrotrophic, and thermoduric microorganism counts were determined by the Standard Plate Count technique. Isolation and identification of E. coli were carried out using conventional physiological tests. Commercial antisera were used for serological characterization of EPEC. RESULTS: The three milk brands analyzed revealed bacterial counts above the regulated values of the Brazilian government. It was found that among 208 strains of E. coli isolated, 46 (22.1%) were serologically classified as EPEC. The most common EPEC serogroup was O55 (15.2%). CONCLUSIONS: Though recent studies on virulence factors indicate that not all strains serologically classified as EPEC are able to attaching/effacing lesion, it is believed that the isolation of EPEC serogroups from pasteurized milk represent a potential risk for children, as well as an indicative of the presence of other enteropathogens.     

Prevalence of enterotoxigenic Escherichia coli-associated diarrhoea and carrier state in the developing world

This study assesses the importance of enterotoxigenic Escherichia coli (ETEC) as a diarrhoeal agent in developing countries. Odds ratios were calculated for incurring ETEC-associated diarrhoea based on data reported between 1970 and 1999. Carriage of ETEC was associated with diarrhoea in children aged less than five years, except for hospitalized infants aged 0-11 month(s) and children aged 1-4 year(s) at outpatient clinics. Two hundred and eighty million episodes of diarrhoea due to ETEC were seen yearly among those aged less than five years, and close to 50 million children of this age group were asymptomatic carriers of ETEC. Every 7th diarrhoeal episode in children aged less than one year and close to 25% of diarrhoeal cases in children aged 1-4 year(s) were due to ETEC. A child born in a developing country is likely to experience 0.5 diarrhoeal episodes per year caused by ETEC until the age of five years, after which the yearly incidence drops to 0.1. To conclude, ETEC remains an important diarrhoeal pathogen among children in the developing world.     

Aetiology of Diarrhoea and Virulence Properties of Diarrhoeagenic Escherichia coli among Patients and Healthy Subjects in Southeast Nigeria

Diarrhoeal diseases are one of the most important causes of illness and death all over the world. In Nigeria, the aetiology of diarrhoeagenic bacteria and the virulence of various Escherichia coli pathotypes have not been well-studied because most currently-published data were from the southwestern axis of the country. In total, 520 stool samples were collected from infants, young children, and other age-groups with acute diarrhoeal diseases in Enugu and Onitsha, southeastern Nigeria. Stool samples were collected from 250 apparently-healthy individuals, with similar age distribution and locality, who were considered control subjects. The stool samples were screened for diarrhea-causing bacterial agents. E. coli strains were isolated from both the groups and were examined by polymerase chain reaction (PCR) for 16 virulence genes. Of the 520 stool samples in the diarrhoea group, 119 (44.74%) were E. coli. Fifty (49.02%) were enteropathogenic E. coli (EPEC), 22 (21.57%) were enterotoxigenic E. coli (ETEC) while 7.84% was enteroaggregative E. coli (EAEC). Sex had no effect on the distribution of diarrhoeagenic bacteria, except for EIEC. The E. coli strains isolated from the diarrhoea and healthy asymptomatic age-matched control groups examined by PCR for 16 virulence genes indicate that the detection of EAEC, ETEC, EPEC, and EIEC was significantly associated with diarrhoea (p=0.0002). The study confirmed that several bacterial pathogens, such as E. coli, play an important role in the aetiology of acute diarrhoea in southeastern Nigeria. A routine surveillance, especially for diarrhoeagenic E. coli, would be useful in identifying outbreaks and help identify the potential reservoirs and transmission routes.Key words: Diarrhoea, Escherichia coli, Virulence, Nigeria     

Characteristic of verotoxigenic strains of Escherichia coli

The aim of the study was the isolation from faecal samples of patients with diarrhoea of verotoxigenic strains of E. coli (VTEC) on the basis of characteristic biochemical properties and production of enterohaemolysin and comparison of isolated verotoxigenic strains with reference strains of VTEC. For isolation of VTEC from 257 stool samples derived from patients with diarrhoea were used selective medium sorbiol--Mac Conkey agar (SMAC) and media supplemented with unwashed and washed in PBS sheep erythrocytes for detection of haemolysins of E. coli. In all haemolytic and sorbitolo-positive or -negative strains isolated from 93 stool samples were examined the activity of beta-glucuronidase using MUG (4-methylumbelliferyl-beta-D-glukuronid) as a substrate for that enzyme. All isolated haemolytic strains as well as reference VTEC were examined on Vero cell line. Verotoxigenic strains from examined samples were investigated by agglutination assay with antiserum to E. coli O157 and then with antisera to eneropathogenic E. coli (EPEC). After that they were examined with ID GN and ATB GN tests. In 93 (36.2%) examined samples there were haemolytic strains of E. coli which fermented or not sorbitol and were MUG-positive or negative. Only in 2 (0.2%) stool samples there were verotoxigenic strains of E. coli which were sorbiol-positive and MG-positive. Both strains belonged to O26 serotype and were derived from samples of two children with diarrhoea. Isolated verotoxigenic strains of E. coli O26 were susceptible on all tested antibiotics.     

Toxin production and haemagglutination in strains of Escherichia coli from diarrhoea in Brescia, Italy

Two hundred and ninety-nine different strains of Escherichia coli, isolated from 172 patients with diarrhoea and 113 healthy subjects, were examined for enterotoxin, cytotoxin and haemolysin (Hly) production and for mannose-resistant haemagglutination (MRHA) and invasive properties. Three strains proved enterotoxigenic, none enteroinvasive; cytotoxin and Hly production was shown in 25 strains from patients and in 3 from controls. Ten strains produced the cytotoxic necrotizing factor (CNF), 6 released other factors which kill cell cultures. Hly production was shown in 21 strains, 9 of which were also positive for CNF. MRHA was detected in 26% of strains from diarrhoea compared with 14% of strains from healthy people. A strong association between toxin production and MRHA was demonstrated. Serotyping results showed that the strains exhibiting virulence traits mostly belonged to serogroups commonly involved in extra-intestinal infections. The possible role of strains of E. coli showing one or more virulence factors as opportunistic pathogens in diarrhoeal diseases is discussed.     

Toxin production and haemagglutination in strains of Escherichia coli from diarrhoea in Brescia, Italy.

Two hundred and ninety-nine different strains of Escherichia coli, isolated from 172 patients with diarrhoea and 113 healthy subjects, were examined for enterotoxin, cytotoxin and haemolysin (Hly) production and for mannose-resistant haemagglutination (MRHA) and invasive properties. Three strains proved enterotoxigenic, none enteroinvasive; cytotoxin and Hly production was shown in 25 strains from patients and in 3 from controls. Ten strains produced the cytotoxic necrotizing factor (CNF), 6 released other factors which kill cell cultures. Hly production was shown in 21 strains, 9 of which were also positive for CNF. MRHA was detected in 26% of strains from diarrhoea compared with 14% of strains from healthy people. A strong association between toxin production and MRHA was demonstrated. Serotyping results showed that the strains exhibiting virulence traits mostly belonged to serogroups commonly involved in extra-intestinal infections. The possible role of strains of E. coli showing one or more virulence factors as opportunistic pathogens in diarrhoeal diseases is discussed.     

腹泻症候群致泻性大肠埃希氏菌血清型及毒力基因检测

目的：通过对腹泻症候群中致泻大肠埃希菌的血清型和多重PCR检测,了解本地区致泻大肠埃希菌的血清群、型分布规律,所携带毒力基因及相互之间的相关性,为腹泻症候群中致泻大肠埃希菌的检测、鉴定提供科学依据,以提高阳性株的检出率。方法：对本地区2012-2013年腹泻症候群监测医院门诊未使用过抗生素腹泻患者的稀便、水样便、黏脓便或脓血便标本通过増菌、各种选择性培养基筛选出病原菌,后经生化试验、多重PCR试验和血清型分型试验进行鉴定。结果：本次检测共检出65株血清凝集致泻大肠埃希菌,分属EPEC、EIEC、ETEC,以EPEC为主,占83.08%,共8个血清型,其中血清型O55：H59、O128：H67占58.47%;共检出4种相关毒力基因,其中escV 49株（75.38%）。未分血清型菌株27株,检出相关毒力基因共5种,分属EPEC、EIEC、ETEC、EAEC,其中escV 15株（55.56%）。结论：本地区腹泻症候群中血清学阳性致泻大肠埃希菌以EPEC为主,常见血清型为O55：H59、O128：H67,毒力基因为escV、escV＋bfpB、invE、elt。未分血清型致泻大肠埃希菌毒力基因为escV、escV＋bfpB、invE、elt、astA,与已分型菌株携带基因基本一致。腹泻患者的大肠埃希菌检测中,在传统的细菌分离培养、生化试验、血清学鉴定的基础上,有必要进行大肠埃希菌的毒力基因检测,以提高阳性检出率,避免漏检,提高致泻性大肠埃希菌的诊断。     

Typical and atypical enteropathogenic Escherichia coli

Typical and atypical enteropathogenic Escherichia coli (EPEC) strains differ in several characteristics. Typical EPEC, a leading cause of infantile diarrhea in developing countries, is rare in industrialized countries, where atypical EPEC seems to be a more important cause of diarrhea. For typical EPEC, the only reservoir is humans; for atypical EPEC, both animals and humans can be reservoirs. Typical and atypical EPEC also differ in genetic characteristics, serotypes, and virulence properties. Atypical EPEC is more closely related to Shiga toxin-producing E. coli (STEC), and like STEC these strains appear to be emerging pathogens.     

碱性磷酸酶标记寡核甘酸探针检测致病性大肠杆菌成束菌毛基因

用合成的碱性磷酸酶标记27bp寡核酸探针检测178株EPEC和320株其他肠道致病菌，并对EPEC菌株同时进行HEp—2细胞粘附及荧光肌料蛋白染色（FAS）试验。结果显示，探针只与EPEC杂交，EPEC中bfpA基因的携带率为25．28％，不同血清人群之间bf－pA基因的携带率有很大差异。HEP—2细胞粘附试验和FAS的检测结果一致。探针杂交与HEp—2细胞粘附及FAS试验相比，符合率为100％，敏感度和特异度分别为95。5％和100％。表明该探针具有很高的特异性和敏感性，操作简便，对健康无害，可用于EPEC的诊断。关键词致病性大肠杆菌成束菌毛寡核甘…     

Aetiology of diarrhoea in a birth cohort of children aged 0-2 year(s) in rural Mirzapur, Bangladesh

The incidence of aetiology-specific diarrhoea and the pathogenicity of infectious agents in a birth cohort (n=252) in rural Bangladesh were determined. Stool specimens or rectal swabs were collected from diarrhoeal cases over two years and routinely on a monthly basis. Stool samples from children with diarrhoea were compared with stool samples from children without diarrhoea to calculate rates of isolation and pathogenicity of agents. In total, 1750 stool specimens from diarrhoea patients and 5679 stool specimens from children without diarrhoea were tested. An infectious agent was identified in 58% of the stool specimens from diarrhoea patients and 21.6% of the stool specimens from children without diarrhoea. The most commonly-isolated pathogens from all specimens were enterotoxigenic Escherichia coli (ETEC), enteroadherent E. coli, Shigella, Campylobacter jejuni, Giardia, and rotavirus. ETEC (ST and LT-ST toxin), enterotoxigenic Bacteroides fragilis, Shigella, and rotavirus were associated more with disease than with asymptomatic infections. Aetiology-specific infections were associated with acute episodes. The isolated enteropathogens were essentially the same as those found in other tropical rural settings. Enterotoxigenic B. fragilis was also identified as a pathogen. Ongoing vaccine efforts focusing on Shigella, rotavirus, and ETEC would be useful.     

Characterization of Salmonella enterica and detection of the virulence genes specific to diarrheagenic Escherichia coli from poultry carcasses in Ouagadougou, Burkina Faso

One hundred chicken carcasses purchased from three markets selling poultry in Ouagadougou, Burkina Faso, between June 2010 and October 2010 were examined for their microbiological quality. The presence of Salmonella was investigated using standard bacteriological procedures, and the isolates obtained were serotyped and tested for antimicrobial susceptibility. The presence of virulence-associated genes of the five main pathogroups of diarrheagenic Escherichia coli-Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli (EPEC), enteroaggregative E. coli (EAEC), enterotoxigenic E. coli, and enteroinvasive E. coli-was investigated using 16-plex polymerase chain reaction (PCR) on the mixed bacterial cultures from the poultry samples. Of the 100 chicken carcasses studied, 57 were contaminated by Salmonella; 16 different serotypes were identified, the most frequent being Salmonella Derby, found in 28 samples. Four Salmonella strains were resistant to tetracycline, and two were resistant to streptomycin. Based on the PCR detection of the virulence genes, in total, 45 carcasses were contaminated by three pathogroups of E. coli: STEC, EPEC, or EAEC. The STEC and EPEC virulence genes were detected on six and 39 carcasses, respectively. EAEC virulence genes were only detected in combination with those of EPEC (on 11 carcasses) or STEC (on two carcasses). The STEC-positive carcasses contained the genes stx(1), stx(2), eaeA, escV, and ent in different combinations. None of the EPEC-positive carcasses contained the bfp gene, indicating that only atypical EPEC was present. EAEC virulence genes detected were aggR and/or pic. The high proportion of chicken carcasses contaminated by Salmonella and diarrheagenic E. coli indicates a potential food safety risk for consumers and highlights the necessity of public awareness of these pathogens.     

Slide-agglutination for rapid serological typing of Treponema hyodysenteriae

A slide agglutination (SA) test was developed to determine the serogroup of isolates of Treponema hyodysenteriae of serogroups A to F. Rabbit antisera which are normally used for serogrouping T. hyodysenteriae in an agarose gel double-diffusion precipitation test (AGDP) were not suitable for SA because they agglutinated isolates from more than one serogroup. The agglutination reaction was made serogroup-specific by cross-absorbing the typing sera for serogroups A to F with whole treponemes from the other 5 of these 6 serogroups of T. hyodysenteriae. The absorbed sera were reacted in slide agglutination tests with 33 isolates of T. hyodysenteriae and with four non-T. hyodysenteriae intestinal spirochaetes. None of the non-T. hyodysenteriae isolates agglutinated, but 27 of the 33 isolates of T. hyodysenteriae did. The results for 26 of the 27 agglutination reactions agreed with the serogroup as determined in AGDP. One of the 6 isolates of T. hyodysenteriae which failed to react in slide agglutination was of serogroup B, 1 of serogroup D, 1 each were from new serogroups G, H and I, and 1 was untypable in AGDP.     

Applicability of a multiplex PCR to detect the seven major Shiga toxin-producing Escherichia coli based on genes that code for serogroup-specific O-antigens and major virulence factors in cattle feces

An 11-gene multiplex polymerase chain reaction (mPCR) was developed based on genes that code for serogroup-specific O-antigens and four major virulence factors (intimin, enterohemorrhagic hemolysin, and Shiga toxins [Stx] 1 and 2), to detect O157 and the "top six" non-O157 (O26, O45, O103, O111, O121, and O145) Shiga toxin-producing Escherichia coli (STEC). The assay specificity was validated with pure cultures of seven major STEC (185 strains), 26 other STEC (65 strains), non-STEC (five strains), and 33 strains of other genera and species. Sensitivity of the assay with cattle fecal sample spiked with pooled cultures of seven major STEC was 10(5) colony-forming units (CFU)/g before enrichment and 10(2) CFU/g after enrichment. The applicability of the assay to detect STEC in fecal samples (n=50), before and after enrichment, was evaluated by comparing with culture-based methods for O26, O111, and O157. The mPCR assay of 50 fecal samples showed seven (14%) positive before enrichment and 23 (46%) positive after enrichment for one or more of the seven O-groups. Overall, 17 isolates from 17 fecal samples and 27 isolates (four for O26, three for O45, and 20 for O103) from 19 fecal samples were obtained, by culture-based methods, for O157 and non-O157 serogroups, respectively. None of the 27 non-O157 isolates possessed the stx genes, suggesting that cattle harbor Shiga toxin-negative E. coli belonging to the "top six" non-O157 serogroups. Our data, although based on a limited number of samples, suggest that the sensitivities of the mPCR and culture-based methods in detecting the seven serogroups of STEC in feces differed between O-groups. An obvious limitation of our mPCR is that the concurrent detection of virulence genes and the serogroups in a sample does not necessarily associate the virulence genes with the prevalent serogroups in the same sample. The major application of our 11-gene mPCR assay may be in identifying putative colonies of STEC obtained by culture-based methods.