To assess the importance of infection by Verotoxin (VT) producing Escherichia coli (VTEC) in children with HUS in Central Europe, stool and/or serum samples obtained from 147 patients from 28 paediatric centres were prospectively examined for the presence of VTEC and the kinetics of faecal VT titres (FVT), and for VT neutralization titres and antibodies against E. coli O 157 lipopolysaccharide, respectively. Ninety-two percent of the patients had classic (enteropathic) HUS (E+ HUS). Evidence of VTEC infection was obtained in 86% of them. VTEC/FVT were identified in 55/118 E+ cases (47%). A prominent feature was the frequent isolation of sorbitol-fermenting, VT2-producing E. coli O 157. H−. VT1 (C600/H19) was neutralized by 9%, and VT2 (C600/933W) by 99% of the initial serum samples from E+ patients, compared to 3% (VT1) and 100% (VT2) from age-related controls. Fourfold titre rises against VT1 and/or VT2 were observed in 13/70 (19%), and significantly elevated O 157 LPS IgM and/or IgA antibodies in 106/128 (83%) of the E+ patients. The ubiquitous VT2 neutralizing principle in the serum of HUS patients as of healthy controls warrants further investigations.

Genotyping was used to analyse Pseudomonas aeruginosa isolates from sink drains and 15 intubated patients as part of a 3-month prospective study of strain transmission in a medical-surgical intensive care unit. Ninety percent of all washbasin drains were persistently contaminated with several P. aeruginosa genotypes. In 60% (9/15) of the patients, P. aeruginosa colonization or infection was hospital-acquired: P. aeruginosa strains isolated from these patients were present in hospital sinks or in other patients before their admission. Since all patients were immobile, personnel were the probable route of transmission of P. aeruginosa in the hospital. The mechanism of strain transmission from sinks to hands during hand washing was investigated in a children's hospital. When P. aeruginosa was present at densities of > 105/c.f.u. per ml in sink drains, hand washing resulted in hand contamination with P. aeruginosa via aerosol generation in the majority of experiments or P. aeruginosa was detected using an air sampler above the washing basin. High P. aeruginosa cfu were present at 4.30 h in the eight sinks (5.4 × 105−7.0 × 1010 c.f.u./ml), whereas at 13.00 h P. aeruginosa c.f.u. were significantly lower (3.1 × 102−8.0 × 105 c.f.u. / ml). These data reveal that the danger of bacterial contamination of hands during hand washing is highest in the morning. The identified transmission routes demand more effective hygienic measures in hospital settings particularly concerning personnel hands and sink drains.

Haemolytic uraemic syndrome (HUS) is a leading cause of acute renal failure in childhood. Although infection with Escherichia coli O 157. H7 has been associated with HUS in North America and Europe, only a limited number of studies have examined the role of other verotoxin-producing E. coli (VTEC) serotypes in this condition. To address this issue, we conducted a comprehensive, prospective microbiological study of patients treated for HUS at eight Canadian hospitals in the summer of 1990. Of the 34 consecutive patients with HUS enrolled over 4 months, E. coli 0 157. H7 was isolated from the stools of 26, and other E. coli serotypes were isolated from four patients. In four subjects no pathogenic E. coli serotypes were identified on stool culture. Using oligonucleotide probes specific for VT-1 and VT-2, verotoxin genes were detected in the stool isolates of all patients with E. coli O 157.H7, and from two with other E. coli serotypes. Two other patients had at least a fourfold rise in anti-verotoxin antibodies. Strong evidence of exposure to a verotoxin was present in 30/34 (88%). Patients with E. coli 0 157.H7 infection were more likely to develop an antibody response to VT-2 than to VT-1 (22/22 vs 12/22; P = 0.002). These results further strengthen the association of HUS with verotoxin-producing E. coli in North America, and confirm that E. coli serotypes other than 0 157. H7 are isolated in a small proportion of summertime HUS episodes.

Two cohorts of 10 and 16 calves were followed at weekly or fortnightly intervals from 4–28 and 1–9 weeks respectively to determine whether natural infection by Vero cytotoxin (VT) producing Escherichia coli (VTEC) occurred. Ninety-one of 171 (53%) faecal specimens were VTEC positive and 20–80% of animals at any given time excreted VTEC. Of 104 VTEC strains studied further, 6 different serogroups (O 22.H16; O 25.H5; O 49.H−; O 86.H26; O 88.H25; O 153.H12) and an untypable strain (O ?.H21) were identified. All strains belonging to the same serotype had identical profiles of reactivity with DNA probes to toxins VT1 or 2, LT1 or iI and a probe (CVD419) derived from a plasmid carried by enterohaemorrhagic Escherichia coli O 157.H7. Four of these serotypes were found in the faecal flora of the calves, taken as a group, throughout the 4-month study period. Sixty percent of the strains hybridized with the probe for VT1, 4% with the probe for VT2, and 36% with both probes. Faecal VTEC were significantly associated with overt diarrhoeal illness in animals < 10 weeks of age, but no characteristic profile of markers (serotype or hybridization pattern) in E. coli isolates was associated with diarrhoea. A serological response to VTl was detected in some animals, but faecal VT1 VTEC excretion persisted in spite of seroconversion. VTl seroconversion was not associated with diarrhoea. A serological response to VT2 was not detected even in those animals excreting VT2 VTEC in the faeces.

One hundred and two Shigella spp. isolated in two hospitals in Hong Kong were analysed for antibiotic resistances, resistance plasmids and plasmid profiles. Three quarters of the isolates were S.flexneri. All isolates harboured plasmids, up to a maximum of ten within one strain. Plasmids of 220 kb encoding resistances to tetracycline, chloramphenicol and sulphonamide and probably also associated with invasivenes in the Sereny test were found in 80 strains and were transferable in 18% of cases. Resistance plasmids of 92 and 99 kb were found in 27 and 15 strains respectively and encoded resistances to ampicillin, tetracycline, chloramphenicol, kanamycin, sulphonamide, trimethoprim, cotrimoxazole and gentamicin; these plasmids were usually transferable. Four plasmids of 3·9, 2·8, 2·2 and 1·8 kb were commonly found in S. flexneri strains, but were rare in other species. In contrast, there was no predominant plasmid profile in S. sonnei. S. flexneri is endemic in Hong Kong and these plasmid studies suggest that the strains in circulation are derived from only a few clones.

To determine whether the risk factors for childhood haemolytic uraemic syndrome (HUS) are similar to risk factors previously reported for Escherichia coli O 157. H7 gastroenteritis, we conducted a case-control study at eight paediatric hospitals in the summer of 1990. Thirty-four consecutive children with HUS were prospectively enrolled; all had diarrhoea and 88% had laboratory evidence of exposure to verotoxin-producing E. coli (VTEC). The 102 controls were otherwise healthy children with minor acute injuries. Parents of all subjects responded to a questionnaire about each child's exposure to various foods, methods of food preparation, sources of water, travel, and individuals with diarrhoea.

Children with HUS were significantly more likely than controls to have had close contact with an individual with diarrhoea in the 2 weeks before the onset of illness (74 υ. 29%, P < 0.00001; odds ratio 7·0. 95% CI 2.7–18.5). The onset of diarrhoea in the contacts occurred a median of 6 days (range, 1− > 14 days) before the onset of diarrhoea in the HUS patients. Exposure to undercooked ground meat was not significantly more common in the patients with HUS (15 υ. 8%; P=0.05). These data provide evidence consistent with person-to-person transmission of VTEC in a substantial proportion of episodes of childhood HUS.

Plasmids in selected type strains of 26 of the Salmonella enteritidis phage types have been characterized by restriction enzyme fingerprinting and by DNA–DNA hybridization with oligonucleotide probes for Salmonella plasmid virulence (Spv) genes. With one exception, the fingerprints of the 38 MDa plasmids studied were homogeneous but there was heterogeneity in the fingerprints of 59 MDa plasmids found in 4 of the type strains. However all 38 MDa and 59 MDa plasmids were related as was a 45 MDa plasmid identified in the type strain of phage type 19. A 3·5 kb fragment homologous to SpvC was conserved in Hind III digests of all 38 MDa and 59 MDa plasmids, and in the related 45 MDa plasmid. In contrast a 65 MDa plasmid found in the type strain of phage type 10 was not related to these three plasmid molecular weight groups and did not carry the SpvC gene.

The present study was undertaken to investigate the occurrence of antibiotic resistance in enteric flora in 64 children in rural Bangladesh over a 12-month period. The antibiotic resistance pattern of the isolates varied throughout the year and multiple resistance was highest during the post monsoon period. Seventythree percent of children had isolates resistant to more than three antibiotics throughout the year. Resistance to streptomycin was highest (78%), followed closely by ampicillin (72%). Of 82 multiply resistant isolates, plasmid DNA was demonstrated in 75%. Plasmid sizes ranged between 3·7 and 110 MDa, the commonest plasmids were of 70, 98 and 110 MDa. Complete or partial resistance was transferred by conjugation from 52% of the isolates, most frequently by single plasmids. The commonest plasmid incompatibility group was F11-A (46%) followed by incompatibility group P (22%). Plasmids of molecular weight 98 MDa most often hybridized with F11-A probes and those of 110 MDa with H11 probes. Plasmids from 10 transconjugants were digested with restriction enzymes and digest patterns demonstrated the presence of common plasmids. The findings show that there is a diverse, and mobile, genetic pool of resistance genes in this rural community. This genetic reservoir is potentially transferable to enteric pathogens, with major implications for public health and diarrhoeal disease control.

Infections due to Salmonella enteritidis are increasing worldwide. In the United States, between 1985 and 1989. 78% of the S. enteritidis outbreaks in which a food vehicle was identified implicated a food containing raw or lightly cooked shell eggs.

Under a US Department of Agriculture regulation published in 1990, eggs implicated in human food-borne S. enteritidis outbreaks were traced back to the source flock. The flock environment and the internal organs of a sample of hens were tested for S. enteritidis. We compared the S. enteritidis phage types of isolates from 18 human, egg-associated outbreaks and the 15 flocks implicated through traceback of these outbreaks. The predominant human outbreak phage type was recovered from the environment in 100% of implicated flocks and from the internal organs of hens in 88% of implicated flocks we tested. The results support the use of phage typing as a tool to identify flocks involved in human S. enteritidis outbreaks.

An analysis of genotype was made for representative strains of Salmonella dublin. The collection consisted primarily of strains isolated from humans in England and Wales, and were of both intestinal and extra-intestinal origin. Three genetic elements were characterized by DNA hybridization. They were the sprBC genes, extrachromosomal virulence determinants. the salmonella-specific insertion sequence IS200, and the 16S ribosomal RNA genes, a phylogenetic marker. Two clones of S. dublin (SdRI and SdRII) which shared an identical IS200 profile, were identified on the basis of restriction fragment length polymorphism at the 16S rRNA locus. With one exception, all strains harboured a 52 MDa plasmid which contained a conserved 3·7 kbp Hind III fragment homologous to the sprBC mouse-virulence genes of S. typhimurium. However, a single plasmid-free strain of SdRI, isolated from a patient with septicaemia exhibited no spc homology. In SdRI there was no observable genotype distinction between strains causing gastroenteritis or bacteraemia. In contrast, none of the strains of SdRII were from cases of bacteraemia, and all human isolates of this clone were from cases of gastroenteritis.

Shigella sonnei is a major agent of diarrhoeal disease in developed as well as in developing countries. Several phenotypic methods to define strain differences have been applied to this species of Shigella including, more recently, analysis of extrachromosomal and chromosomal DNA.

In this study, 432 endemic and epidemic strains isolated between 1975 and 1991 in Italy, France and Switzerland were submitted to rRNA gene restriction pattern analysis, after digestion of whole-cell DNA by Hinc II, and to concomitant biotyping.

Thirteen ribotypes, HI to H13, and five biotypes, a, d, e, f, g, were detected. Xinety-five percent of the sporadic strains were assigned to ribotypes HI to H4, which could be subtvped, except for H4, in different biotypes. Strains from each of seven different outbreaks had indistinguishable ribotype-biotype patterns. In contrast, 65 strains, isolated in Sicily in 1980 over an extended period of apparently epidemic increase of isolations and which had previously been considered to be a single bacterial clone on the basis of resistance pattern and phage type, were found to belong to two different and scarcely related ribotypes.

Ribotyping and biochemical subtyping appear to be a useful epidemiological tool in studies on the circulation and distribution of strains of S. sonnei.

Plasmids belonging to the FIme incompatibility group were found in seven different serogroups of multiply antimicrobial-resistant Escherichia coli isolated from patients with urinary tract infection (UTI) and living in south-east London. Although widespread in Salmonella spp., FIme plasmids have only previously been described in E. coli in a strain of serogroup O15 K52 H1 responsible for an extensive and protracted outbreak of invasive community-acquired infection in south-east London in 1986. Our findings suggest either a wider background occurrence of FIme plasmids in E. coli associated with UTI than previously reported or alternatively, the dissemination and subsequent molecular diversification of the FIme plasmid associated with the epidemic strain of serogroup O15 K52 H1.

Between 1989 and 1991 a total of 1092 Vero cytotoxin-producing Escherichia coli O 157 isolated in the United Kingdom were phage typed in the Laboratory of Enteric Pathogens (LEP). Twenty-three phage types was identified, the most frequent being types 2 (36·1%), 49 (29·6%), 1 (10·3%) and 4 (8·9%). Although isolations of O 157 VTEC have increased each year from 1 in 1982 to 532 in 1991, the predominant phage types have remained unchanged although the proportion of strains belonging to types 2 and 49 has increased. O 157 VTEC from 17 outbreaks were phage typed during this period with phage type 49 predominating (7 of 17 outbreaks).

In recent years, Salmonella enteritidis has become an increasingly important public health problem in Italy. In some parts of the country, the fraction of total human salmonella isolates accounted for by S. enteritidis has risen from 3–4% in the mid-1980s to more than 30% in 1990. Between 1090 and 1991. the number of reported S. enteritidis outbreaks increased more than sixfold. The 33 outbreaks reported in 1991 occurred in seven contiguous regions in northern and central Italy and were clustered in time between June and October: in the majority, products containing raw or undercooked shell eggs were implicated. Five of the egg-related outbreaks that occurred within a 30 kilometre radius over a 7-week period were investigated in detail. A phage type 1 strain containing a 38·9 MDa plasmid appeared responsible for three of the outbreaks, while in the remaining two a phage type 4 strain, also with a 38·9 MDa plasmid was isolated. Efforts are being made to enhance epidemiological surveillance and laboratory evaluation, and the use of pasteurized eggs has been recommended for high-risk populations.

Between January and June 1990, Restaurant A in Greenville, South Carolina repeatedly failed local health department inspection and was repeatedly sanctioned. In September 1990, two persons, hospitalized with salmonellosis after attending a convention catered by Restaurant A, contacted the South Carolina Department of Health and Environmental Control. We inspected Restaurant A, interviewed food handlers, and surveyed by telephone persons from every sixth business attending the convention. Of 398 persons interviewed, 135 (34%) reported gastroenteritis. Nine had culture-confirmed salmonella infection. People who ate turkey were 4.6 times more likely to become ill than those who did not eat turkey (95% confidence interval 2.0, 10.6). We estimate that of 2430 attendees, 824 became ill. Sanitarians judged Restaurant A's kitchen too small to prepare over 500 meals safely. The cooked turkey was unrefrigerated for several hours, incompletely rewarmed, and rinsed with water to reduce its offensive odour prior to serving. Stronger sanctions may be needed against restaurants that repeatedly fail local health department inspection.

An outbreak of Salmonella typhimurinm definitive type (DT) 193 affecting 206 persons occurred in July and August 1989 in a small town in northern England. A descriptive study suggested that cold meats including pork from a butcher's shop in the town were vehicles of infection. An analytical study of a cohort attending a function in the town showed a significant association between illness and consumption of cold roast pork supplied by the butcher's shop (P = 0·00000004). S. typhimurium DT 193 with the same antibiotic resistance pattern (to ampicillin, streptomycin, sulphonamides and tetracyclines) as the outbreak strain, and possessing a single plasmid of 80 MDa was isolated from samples of meat bought from the shop and implicated in illness, and from samples of pig faeces taken from the farm supplying the shop. It was concluded that inadequate processing of infected pork meat at the shop may have contributed to this outbreak but that cross contamination also played an important part in transmission. Control measures included a temporary closure of the shop and subsequent implementation of a detailed protocol for meat processing and monitoring of all procedures at the shop.

Strains of Salmonella enteritidis were examined for their ability to remove ferricions from the iron chelating agents ovotransferrin, Desferal and EDDA. Growth of S. enteritidis phage type (PT) 4 (SE4) in trypticase soy broth containing ovotransferrin resulted in the expression of iron regulated outer membrane proteins (OMPs) of 74. 78 and 81 kDa. and unexpectedly the repression of expression of OMP C. The 38 MDa ‘mouse virulence’ plasmid was not required for the expression of the iron-regulated OMPs (IROMPs). SE4 was able to obtain iron bound to the iron chelator Desferal and EDDA without expressing a high-affinity iron uptake system. Strains of S. enteritidis belonging to PTs 7. 8, 13a, 23. 24 and 30 were also able to remove ferric ions from Desferal and EDDA without expressing a high-affinity iron uptake system. We conclude that strains of SE4 possess a high-affinity iron sequestering mechanism that can readily remove iron from ovotransferrin. It is likely that iron limitation, and not iron restriction, is responsible for the bacteriostatic properties of fresh egg whites.

Over the 7 years 1985–91, 997 strains of Staphylococcus aureus from 962 patients with diseases other than food poisoning have been tested for the production of enterotoxins and toxic shock syndrome toxin-1 (TSST-1) and phage typed. In all, 128 cases could be classified as confirmed or probable toxic shock syndrome (TSS) but a further 199 cases were classified as possible or unconfirmed TSS. In 219 cases, an alternative diagnosis could be supported and 45 cases were classified as sudden infant death syndrome. In 371 cases, insufficient information for classification was available.

Strains of phage group I producing TSST-1 were associated with menstrual TSS. Many menstrual TSS cases were aged less than 20 and were using non-introducer tampons.

When all strains were reviewed, strong associations were observed between TSST-1 production and phage group I strains, enterotoxin B production and group V strains, enterotoxin C and phage-type 95 strains and between enterotoxin A without TSST-1 and phage group III strains.

In 1990, a Salmonella enteritidis (SE) outbreak occurred in a restaurant chain in Pennsylvania. To determine its cause(s), we conducted a case-control study and a cohort study at one restaurant, and a survey of restaurants. Egg dishes were associated with illness (P = 0.03). Guests from one hotel eating at the restaurant had a diarrhocal attack rate of 14%, 4.7-fold higher than among those not eating there (P = 0.04). There were no differences in egg handling between affected and unaffected restaurants. Eggs supplied to affected restaurants were medium grade AA eggs from a single farm, and were reportedly refrigerated during distribution. Human and hen SE isolates were phage type 8 and had similar plasmid profiles and antibiograms. We estimate the prevalence of infected eggs during the outbreak to be as high as 1 in 12. Typical restaurant egg-handling practices and refrigeration during distribution appear to be insufficient by themselves to prevent similar outbreaks.

Cholera vibrios can be allocated to one of three biotypes (classical, intermediate and El Tor), each of which can be sub-divided into two serotypes (Ogawa and Inaba). Vibriocidal tests with absorbed antisera have shown no evidence of biotype specificity in the killing of bacteria, but they have confirmed the role of the two serotype-specific antigens in immunity to cholera. The same presence of serotype specificity, and absence of biotype specificity, has been found by bacterial agglutination, an easier and quicker serological test. The use of this simpler test in ensuring a balanced serotype response to cholera vaccine is discussed, together with evidence that may lead to the production of more effective vaccine and better immunity.