Sylvia M. Scotland

First recognized community outbreak of haemorrhagic colitis due to verotoxin-producing Escherichia coli O 157.H7 in the UK

The first recognized outbreak of haemorrhagic colitis due to Escherichia coli O 157.H7 in the United Kingdom affected at least 24 persons living in East Anglia over a 2-week period. The illnesses were characterized by severe abdominal pain and bloody diarrhoea of short duration. Eleven patients were admitted to hospital and there was one death. Patients were mainly adult women who had not eaten out of the home in the 2 weeks before onset. Unlike previously reported outbreaks hamburgers were not the vehicle of infection, and a case-control study suggested that handling vegetables, and particularly potatoes, was the important risk factor.

Surveys of human enterotoxigenic Escherichia coli from three different geographical areas for possible colonization factors

Enterotoxigenic Escherichia coli (ETEC) from Burma, central Africa (Rwanda and Zaire) and Peru, were screened by enzyme-linked immunoassays for the colonization factor antigens (CFAs) and putative colonization factors (PCFs): CFA/I, CFA/II, which consists of three coli surface-associated (CS) antigens, CS1, CS2 and CS3, CFA/III, CFA/IV (CS4, CS5, CS6), CS7, PCFO9, PCFO159. H4, PCFO166, and CS17. The highest proportion of ETEC with identifiable colonization factors (71%) were found in the strains from Burma, which were mainly positive for CFA/I (38%), but strains producing CFA/II (4%), CFA/IV (11%), CS7 (10%), CS17 (4%), PCFO159, H4 (2%) and PCFO166 (2%) were also found. Sixty-nine percent of the ETEC from central Africa were positive for known colonization factors. While CFA/I positive strains were important (12%), a higher number of ETEC producing CFA/IV (33%) and CS17 (24%) were found. Fifty-two percent of the Peruvian strains produced identifiable colonization factors. The largest group of strains produced antigens of the CFA/IV complex (17%), while ETEC producing CFA/II (6%), CFA/III and CS6 (2%), CS7 (6%), PCFO9 (6%), PCFO166 (8%) and CS17 (7%) were also found. These surveys show that there is a considerable variation in the proportions and types of colonization factor found in different geographical areas. From 29 to 48% of the ETEC did not possess an identifiable colonization factor. These were particularly of the LT only producing type. These results have important implications for vaccine formulation.

HEp-2 Adhesion and the Expression of a 94 kDa Outer-membrane Protein by Strains of Escherichia coli Belonging to Enteropathogenic Serogroups

Sixty strains of Escherichia coli belonging to enteropathogenic serogroups (EPEC) were examined for the ability to adhere to HEp-2 cells, the possession of the genes encoding EPEC adherence factor (EAF) and the ability to express an outer-membrane protein (OMP) of 94 kDa thought to be involved in bacterial adhesion to eukaryotic cells. An absolute correlation was found between HEp-2 adhesion and the possession of the genes encoding EAF. An OMP of 94 kDa was observed in the SDS-PAGE profile of most adhesive strains. In some strains this protein was prone to proteolytic degradation. An antiserum raised to a HEp-2 adhesive strain of EPEC did not react with the 94 kDa OMP of all EPEC which were EAF-positive and HEp-2 adhesive, indicating some interstrain antigenic variation of this protein. Although this 94 kDa protein was surface-exposed, specific antibodies binding to the 94 kDa protein in situ in the outer membrane did not interfere with adhesion of EPEC to HEp-2 cells. Therefore, these studies question the value of this protein as a potential vaccine component.

A method of enhancing verocytotoxin production by Escherichia coli

The number of verocytotoxin producing Escherichia coli (VTEC) present in the faeces during an infection may be very low, making their detection difficult. We report a method for enhancing toxin production by VTEC using mitomycin C as an inducing agent with the aim of improving the detection of VTEC. In pure culture, mitomycin C enhanced toxin production up to 100-fold. When applied to mixed faecal culture, toxin could be detected in mitomycin C treated samples when standard cultures were negative and when substantially fewer verocytotoxin-producing bacteria were present. Use of this method may aid in the detection of VTEC and is appropriate for use in the routine diagnostic laboratory.

Properties of adherence factor plasmids of enteropathogenic Escherichia coli and the effect of host strain on expression of adherence to HEp-2 cells.

EPEC adherence factor (EAF) plasmids from three strains of enteropathogenic Escherichia coli (EPEC) - E2347/69 (O127:H6), E20517 (O111:H2) and E24582 (O142:H6) - were examined. The EAF plasmids were all marked with ampicillin resistance by transposition of Tn801 to give pDEP1, pDEP2 and pDEP11, respectively. All three plasmids showed incompatibility with an FIme and an FIV plasmid and had some similarity in restriction enzyme digest patterns. Plasmid pDEP1 differed from pDEP2 and pDEP11 in being autotransferring and fertility-inhibition positive. An EAF probe consisting of a 1 kb BamHI-SalI restriction endonuclease fragment of the prototype EAF-associated plasmid pMAR2 hybridized to similar-sized SalI-BamHI fragments of pDEP1 and pDEP11 but to a different-sized fragment of plasmid pDEP2. Loss of the EAF plasmids from EPEC strains resulted in a marked reduction in the ability of these strains to adhere to HEp-2 cells. The EAF-plasmid-negative variants did not express a 94 kDa outer-membrane protein (OMP). When these EAF plasmids were reintroduced into EAF-plasmid-negative EPEC strains a high level of adherence equivalent to that of the parent EPEC strains was restored and a 94 kDa OMP was usually expressed. However, when EAF plasmids were transferred into E. coli K12 or non-EPEC E. coli the host strains either did not adhere or adhered poorly to the HEp-2 cells. These transconjugants did not express a 94 kDa OMP.

The occurrence of plasmids carrying genes for both enterotoxin production and drug resistance in Escherichia coli of human origin.

Twenty-three of 89 enterotoxigenic strains of Escherichia coli were resistant to one or more antimicrobial agents. Eleven strains transferred resistance directly and five transferred resistance after mobilization. In three cases a resistant recipient was enterotoxigenic. One of these strains contained a conjugative plasmid carrying genes for both drug resistance and enterotoxin production. In the two other strains genes for drug resistance and enterotoxin production were carried on separate co-transferable plasmids.

Serotype-related enterotoxigenicity in Escherichia coli O6.H16 and O148.H28.

The ability of certain Escherichia coli strains to produce enterotoxin is determined by transmissible plasmids. It is therefore possible that any E. coli strain might be able to acquire such a plasmid and that the correlation between enterotoxigenicity and serotype might be random. However, recent studies show that the enterotoxigenic strains so far describe belong to a restricted range of serotypes. Enterotoxigenic strains of E. coli O6.H16 and E. coli O148.H28 have been associated with outbreaks of diarrhoea in several countries, therefor strains of E. coli belonging to these serotypes were selected for further study. Twenty-three strains of E. coli O6.H16 and 14 strains of E. coli O148.H28 were examined; 20 strains of E. coli O6.H16 and all 14 strains of E. coli O148.H28 were enterotoxigenic but strains of E. coli O6 wit flagellar antigens other than H16 and strains of E. coli O148 wit flagellar antigens other than H28 were not enterotoxigenic. The examination of single colony subcultures derived from the E. coli O6.H16 strains showed that in some strains loss of enterotoxigenicity had occurred in a proportional of colonies.

The use of gene probes, immunoassays and tissue culture for the detection of toxin in Vibrio cholerae non-O1

Vibrio cholerae non-O1 strains were screened for the presence of cholera enterotoxin (CT) genes by means of digoxigenin-labelled polynucleotide CTA and CTB probes. In-vitro production of CT was investigated by the Y1 mouse adrenal cell assay, enzyme-linked immunosorbent assay (ELISA) and a commercial, reversed passive latex agglutination (RPLA) kit. Only two (0.25%) of 790 strains tested gave positive results with the CTA and CTB probes. The production of other bacterial cytotoxin(s) made it impossible to use the characteristic cell-rounding effect on Y1 cells for the detection of CT. CT production by the probe-positive strains was confirmed by the immunoassays. Two hundred and fifty-two of the 788 probe-negative strains were tested by both cell assay and immunoassays. Of these, 90% produced cytotoxin(s) in the cell assay. In addition, 37% gave positive results in CT-ELISA, but negative results with LT-ELISA and VET-RPLA. These results indicate the presumed presence of a toxin in V. cholerae non-O1 that is able to bind GM1 and react with antisera to CT, but which is not identical to CT.

Acquisition and maintenance of enterotoxin plasmids in wild-type strains of Escherichia coli

Enterotoxigenic strains of Escherichia coli (ETEC) may produce a heat-labile enterotoxin (LT), a heat-stable enterotoxin (ST) or both enterotoxins. Certain serogroups are represented more frequently than others in ETEC isolated from humans. The transfer of three plasmids encoding enterotoxin production (Ent) to 22 non-toxigenic E. coli strains of many different O:H serotypes was studied. The Ent plasmids encoded ST (TP276), or LT (TP277), or ST + LT (TP214), and all carried antibiotic-resistance determinants. Twenty-one recipient strains acquired TP214, 18 acquired TP277 and 14 acquired TP276. Strains of those serotypes to which ETEC in diarrhoeal studies commonly belong neither acquired nor maintained Ent plasmids with a higher frequency than strains of those serotypes to which ETEC rarely belong. The recipient strains, with one exception, all expressed ST, or LT, or ST and LT, when they had acquired the appropriate plasmid; a non-motile strain belonging to O serogroup 88 expressed LT but failed to express ST when it acquired TP214 or TP277.

The possession of coli surface antigen CS6 by enterotoxigenicEscherichia coli of serogroups O25, O27, O148, and O159: a possible colonization factor?

A total of 134 enterotoxigenicEscherichia coli (ETEC) of serogroups O25, O27, O148, and O159 were tested in the enzyme-linked immunosorbent assays for the colonization factor antigens I (CFA/I), CFA/II (coli surface antigens CS1, 2 and 3) and putative colonization factor (PCF) 8775 (CS4, 5 and 6). CS6 was detected without CS4 or CS5 in 94% of the strains of serogroup O25, 86% of strains of serogroup O27, 87% of strains of serogroup O148, and 29% of strains of serogroup, O159. The frequency with which CS6 occurs in ETEC of common serotypes without the antigens CS4 or CS5 suggests that it might be a colonization factor.