In a setting where potable water is contaminated with Legionella pneumophila serogroup 1, we performed two case control studies. The first case control study consisted of 17 cases of nosocomial Legionnaires' disease (LD) and 33 control (the patients who were admitted to the ward where the case was admitted immediately before and after the case) subjects. Cases had a higher mortality rate 65% vs 12% (P < 0.004); were more likely to have received assisted ventilation (P < 0.00001); to have nasogastric tubes (P < 0.0004) and to be receiving corticosteroids or other immunosuppressive therapy (P < 0.0001). Based on the results of this study, sterile water was used to flush nasogastric tubes and to dilute nasogastric feeds. Only 3 cases of nosocomial LD occurred during the next year compared with 12 the previous year (P < 0.0001). Nine cases subsequently occurred and formed the basis for the second case-control study. Eighteen control subjects were those patients admitted to the same unit where the case developed LD, immediately before and after the case. The mortality rate for the cases was 89% vs 6% for controls (P < 0.00003). The only other significant difference was that cases were more likely to be receiving corticosteroids or other immunosuppressive therapy 89% vs 39% ( < 0.01). We hypothesized that microaspiration of contaminated potable water by immunocompromised patients was a risk factor for nosocomial Legionnaires' disease. From 17 March 1989 onwards such patients were given only sterile potable water. Only two cases of nosocomial LD occurred from June 1989 to September 1990 and both occurred on units where the sterile water policy was not in effect. We conclude that aspiration of contaminated potable water is a possible route for acquisition of nosocomial LD in our hospital and that provision of sterile potable water to high risk patients (those who are receiving corticosteroids or other immunosuppressive drugs; organ transplant recipients or hospitalized in an intensive care unit) should be mandatory.

The morphology of faecal microflora of nine healthy human volunteers was studied by digital image analysis of microscopic slides. Weekly specimens were collected during an 8-week period. Seven morphometrical parameters were derived: the means and medians of components 1, 2 and 3, and morphometrical entropy. Statistically significant differences among subjects were found for means of components 1 and 2, medians of components 1, 2 and 3, and entropy. The stability in normal circumstances provides an excellent basis for the detection of pathological change in gut flora balance.

Following a case of Legionnaires' disease Legionella pneumophila of the same serogroup was isolated from the hot water system of the hotel which had been epidemiologically implicated as the source of the infection. Pyrolysis mass spectrometry (PyMS) was used to compare 11 isolates from the hotel water sample with the patient's strain. Epidemiologically unrelated isolates, both clinical and environmental, of the same serogroup and monoclonal antibody type of L. pneumophila were included in the same analysis, together with relevant reference strains. The patient strain was shown to be indistinguishable from seven of the hotel water isolates, but clearly different from other unrelated clinical isolates, the reference strains and some of the other environmental isolates. PyMS is a rapid and cheap method for inter-strain comparison for L. pneumophila.

The relationship between visual inspections carried out by environmental health officers and microbiological examination was studied in 89 restaurants. Using 30 variables a standardized inspection procedure was developed and each of the premises was assessed in six main areas – structure and design, cleaning and cleanliness, personal hygiene, risk of contamination, temperature control, and training and knowledge about food hygiene. Selected foods and specimens from hands, surfaces, and wiping cloths were examined. There were significant associations between all six areas of the inspections. The structure and design were significantly related to the combined score from all the other areas (P < 0·001). There were no highly significant associations between microbiological examination and visual assessments. The microbial contamination of wiping cloths, however, was related to the cleaning and cleanliness (P = 0·005). Microbial sampling provided additional information to inspections and was a valuable aid. Further development of this risk-assessment approach could provide an effective system for monitoring potential health risks in high-risk food premises.

Twenty-three pre- and post-treatment isolates of Helicobacter pylori from the antral mucosa of eight patients with dyspepsia and gastritis were compared using 1-D SDS PAGE of proteins. The protein patterns were highly reproducible and were used as the basis for two numerical analyses. The first, based on the total protein patterns, showed that a number of the strains did not cluster with their respective patient set. This was thought to be due to differences in both mobility and intensity of proteins in the major band region. The second analysis, based on partial patterns, excluding the major band region (51–68 kDa), divided the clinical isolates into clearly defined groups corresponding to the patient sets. Although there was a degree of heterogeneity with respect to protein pattern between the pre- and post-treatment isolates of some patients, there was nonetheless clear evidence that each patient was harbouring strains of only a single type. These results suggested that patients were not being reinfected with a different strain but that there was recrudescence of the pre-treatment strain. Protein ‘fingerprints’ provided a precise and reproducible means of strain differentiation, and revealed that in each patient the same strain persisted after drug therapy even though there was marked patient-to-patient strain variation.

Fourteen people living in or near the city of Gloucester fell ill with Legionnaires' disease caused by Legionella pneumophila serogroup (SG) 1 between 27 August and 27 October 1986. Another patient had fallen ill on 30 May. Nine of the 15 were diagnosed retrospectively during a case finding exercise. There were three deaths. Three cases of Pontiac fever were also diagnosed.

The source was probably one or more wet cooling towers. Nineteen premises in the city with such towers were identified, and three just outside Gloucester. Samples from 11 of the 22 premises grew Legionella spp.; from nine of these L. pneumophila SG 1 (Pontiac) was isolated. The efficacy of regular addition of biocide in addition to hypochlorite added at the time of disinfection in inhibiting the growth of Legionella spp. was demonstrated.

A survey of patients' movements during their likely incubation period showed that there was no single building that all patients had visited, but there were two areas of the city which nearly all had visited or passed through by car. A case-control study demonstrated an association with one of these areas.

Cooling towers near both areas may have been sources but the evidence is insufficient to incriminate any single one. The unexpected finding of L. pneumophila SG 1 (Pontiac) in nine towers supports the hypothesis that there may have been multiple sources. Cooling towers may have been contaminated by mains water or by drift from other towers.

We have developed a nucleic acid dot-blot hybridization test for the detection of astroviruses in stool samples. The test was not as sensitive as electron microscopy for the detection of low numbers of well preserved astrovirus particles, but was able to identify astroviruses in stools containing particles of indistinct morphology. In total, this procedure identified astroviruses in more samples than did electron microscopy, and the data indicate that the incidence of astroviruses may be substantially underestimated.

The colonization resistance (CR) of the digestive tract was determined in 10 healthy volunteers by oral contamination with a neomycin resistant Escherichia coli (NR-E. coli) strain and measurement of the faecal concentration of this strain during 14 days after the contamination. This ‘gold standard’ was compared with another parameter of CR; the determination of the mean number of different biotypes of Enterobacteriaceae isolated from four faecal samples per volunteer. Both measures are significantly correlated (P < 0.01). The NR-E. coli strain could be cultured from faecal samples of 4/10 volunteers as long as 300 days after contamination. Serum antibody titres against endogenous E. coli strains and the NR-E. coli strain used for experimental oral contamination were measured by an indirect immunofluorescence (IIF) assay. The assay was read by a video camera connected to an image processing system. The 95% confidence limits of antibody titres (log2) against endogenous E. coli strains ranged between < 3 and 7.1 for IgA, between < 3 and 8.7 for IgG and between < 3 and 7.4 for IgM. Antibody titres against the NR-E. coli 4 strain were within this (normal) range. The serum antibody titres against the NR-E. coli strain increased slowly after oral contamination, especially IgG and IgM. Little increase in IgA titres could be observed. An increase of serum antibody titres did not correlate with the elimination of the oral contaminant from the intestines. Therefore, we conclude that the CR is not IgG nor IgM antibody mediated.

Serum samples obtained from patients hospitalized in Barbados with severe leptospirosis were tested by the microscopic agglutination test (MAT), enzyme immunoassay (EIA) and immunoblotting with leptospires that had been isolated from these patients. While serum samples taken a few days after onset of symptoms often showed no apparent correlation between MAT and EIA, later sequential serum samples produced similar profiles in both tests during the course of infection. Immunoblotting sonicate from Leptospira interrogans serovars arborea, copenhageni and bim with patients' sera, revealed reactions with a number of bands that corresponded with outer envelope components. These components included lipopolysaccharide (LPS), flagella and other outer membrane proteins, in addition to a low-molecular-weight (MW) carbohydrate cross-reactive with members of the Leptospiraceae. IgM antibodies elicited in the first to second week after infection reacted mainly with LPS and the low-MW cross-reactive carbohydrate. Comparative analysis of isolates of the same serovar by sodium dodecyl sulphate polyacrylamide gel electrophoresis and immunoblotting showed that while two serovar arborea isolates were identical, serovar bim isolates differed significantly from each other. This difference was also observed in comparative MAT testing.

Relapse of Clostridium, difficile-associated diarrhoea occurs in 15–20% of patients; however, whether relapse is due to an endogenous source of the organism or reinfection from the environment remains unclear. Restriction enzyme analysis (REA) of chromosomal DNA was used to type multiple isolates from ten patients who had experienced apparent relapses. More than half the relapses were due to infection with a new strain of C. difficile. The remaining patients were infected with the same strain, but whether this strain was acquired from the environment or from endogenous sources could not be determined. Relapses with a different strain of C. difficile could occur if an individual harboured more than one strain in their gastrointestinal tract. To investigate this possibility ten other patients were assessed for carriage of multiple strains. Ten colonies from a primary culture plate from each patient were typed by REA and tested for their ability to produce cytotoxin. All isolates from the same patient were identical by both methods, indicating that multiple carriage of strains may be a rare event.

Numerous investigators have reported an increased incidence of pneumonia caused by Gram-negative bacilli and other secondary pathogens in transplant recipients infected by cytomegalovirus (CMV). To determine if CMV infections are related to colonization of the upper respiratory tract by Gram-negative bacilli, we examined prospectively 22 renal transplant recipients with sequential bacteriological, virological and biochemical examinations performed just prior to and at various times after transplantation. Only 11% of subjects had Gram-negative bacilli isolated from gargle specimens prior to transplantation, as compared to 54% after transplantation. More importantly, after transplantation, subjects with active CMV infections were more likely to have prolonged oropharyngeal carriage of Gram-negative bacilli than subjects without CMV infections (36% v.. 25%). During active CMV infections, the rate at which Gram-negative bacilli were isolated from gargle specimens rose from 28 to 47%. During culture-positive CMV infections, the isolation rate reached 57% and was significantly different from that of CMV-negative samples (P < 0·01). The increased rate of Gram-negative bacillary isolation from gargle specimens during CMV infections was not a function of type of immunosuppresive agents used, rejection episodes, antibiotic administration, concomitant hepatitis B, Epstein–Barr (EBV) virus, or herpes simplex virus infections, or alterations in salivary fibronectin concentrations.

Between 1987 and 1989, the prevalence of antibody to cytomegalovirus (CMV) was determined, by the ELISA method, in serum samples from 1494 apparently healthy subjects, 3–18 years old. Subjects were selected by a systematic cluster sampling from five geographical areas in Italy. The overall prevalence of antibody was 64·2%, increasing from 54·4% in 4–6-year-olds to 73·3% in subjects 17–18 years old (P < 0·01). Prevalence of antibody was significantly higher in females (P < 0·05) and in subjects residing in the South of Italy (P < 0·01). A significant association was found with sociodemographic factors. Subjects belonging to a household with six or more persons had a 1·5-fold risk (C.I. 95% = 1·11–2·04) and subjects whose fathers had less than 6 years of schooling had a 1·4-fold risk (C.I. 95% = 1·1–1·87) of previous exposure to CMV infection. The high prevalence (74·4%) of young women who are naturally immune when entering childbearing years does not guarantee that there will be a low risk of fetal infection.

A containment laboratory unit for research with aerosols of group 2 pathogenic microorganisms is described. The design criteria are based on current UK guidelines, which imply containment at group 3 level during aerosol production, storage, exposure of animals and sampling. Within the aerosol laboratory, primary containment is provided by a Henderson apparatus operating at a negative pressure to the external environment. Flexible film isolators under negative pressure are used for all hazardous microbiological work, e.g. tissue homogenization, and for housing infected laboratory rodents. A novel feature of the animal isolator is the separate ventilation of each cage, which minimizes the risk of cross-infection by aerosol transmission and ensures a similar environment within each cage. The results of an intentional release of a cloud of non-pathogenic microorganisms are presented to show the effectiveness of the containment barriers. Recommendations are given for the safe operation of a containment unit based upon practical experience.

Although vaccination is widely practiced against infection by Erysipelothrix rhusiopathiae in pigs and turkeys, the protective antigen(s) involved have not been fully characterized or purified to homogeneity. Antigens of E. rhusiopathiae strain T28, serotype 2b, and of FRANKFURT XI, serotype N, in culture supernatant and in extracts made with hot acid, 10 mM NaOH, ultrasound or EDTA were compared by SDS-PAGE and immunoblotting and in a mouse protection test. EDTA and 10 mM NaOH yielded highly protective extracts; culture supernatant was less protective and ultrasonic or hot acid extracts stimulated little or no protection in mice. Protective antisera from swine, horses and mice recognized prominent bands of molecular mass (m.m.) of 66–64 and 40–39 kDa in EDTA and 10 mM NaOH extracts. Mice immunized with preparations of the 66–64 kDa band purified by preparative electrophoresis were protected. Both antigens were trypsin sensitive, contained no detectable polysaccharide, and showed a marked tendency to aggregate in the absence of SDS.

A matched and serologically confirmed case-control study was carried out to investigate the source of an outbreak of acute hepatitis involving 290000 cases in the suburbs of Shanghai, in January 1988. A total of 132 patients with acute hepatitis from six different hospitals were chosen as cases and the same number of control patients without hepatitis were matched for gender, age, admission date and area of residence. Serum specimens from both case and control patients were detected for specific anti-hepatitis A (HA) IgM antibody and a questionnaire was used to investigate probable risk factors related to the outbreak. The positive rate of anti-HA IgM was 98·48% in the case group and only 0·76% in the control, indicating that the infection was caused by HA virus. The results revealed that the source and mode of transmission were due to the consumption of contaminated and inadequately cooked clams (Anadara subcrenata lischke). There was a highly positive dose-response relationship between the odds ratio of contracting HA and the quantity or frequency of clam consumption. The odds ratios of acquiring HA from clams were up to 62·4–63·4 by both group stratification and multiple unconditional logistic regression analyses.

Aeromonas has been recognized as an important enteropathogen, but factors related to its virulence have not been clarified. For most enteric pathogens, attachment is a prerequisite for infection and for the effective delivery of toxins to the intestinal epithelial cells. We examined a total of 273 strains isolated from stool, food and environmental specimens by an assay for mannose-resistant adhesion to INT407 cells in vitro. Seventeen of the 102 faecal isolates were adhesive strains with more than 10 bacteria adhering per cell, while only 2 of the 118 isolates from foods and river water adhered to the cells (P < 0·001). It is possible that the adhesion might serve as a marker for discrimination between the pathogenic and nonpathogenic isolates. The 8 highly adhesive strains with more than 20 adhering organisms per cell were scrutinized for the mechanism of adhesion. No correlation was apparent between the adhesion to INT407 cells and hydrophobicity. It was noted that fucose inhibited the adhesion of four strains as well as haemagglutination by them. Electronmicroscopic studies showed the presence of flexible and curvilinear fimbriae in only 2 of the 8 highly adhesive strains.

The prevalence of antibodies to human T-cell lymphotropic virus type I (HTLV-1) was determined in high-risk groups and normal adults in Egypt. Among 647 individuals tested, 6 (0·9%) were confirmed positive by western blot analysis. These included 2 (0·7%) of 279 drug addicts, 1 (3·3%) of 30 patients with sexually transmitted diseases, and 3 (2·2%) of 133 healthy individuals. Antibody was not detected in 47 blood recipients or 158 prostitutes. There was no correlation between sex or geographical location and HTLV-I infection. Fifty-three of the 647 sera (8%) were initially reactive by ELISA, but only 12 sera were repeatedly reactive. Since only 4 of these repeatedly reactive sera were confirmed by the western blot, the frequency of false positives using the DuPont screening ELISA was 1·2% (8/643). Two additional sera, confirmed positive by western blot, had been reactive, but not repeatedly, by ELISA. In comparison to the prevalence of HTLV-I antibody among risk groups in many parts of the world, the prevalence in Egypt was low.

A study was conducted to isolate and identify microorganisms of medical importance from cockroaches (Blattella germanica) and to ascertain their vector potential in the epidemiology of nosocomial infections. Bacteria, fungi and parasites of medical importance were isolated and identified. Important bacterial pathogens responsible for wound infections, were further studies by antibiograms. One hundred and fifty-eight out of 159 (99·4%) cockroaches collected from hospital (test) and 113 out of 120 (94·2%) cockroaches collected from residential areas (control) were carrying medically important microorganisms (P < 0·05). A significantly higher (P < 0·001) number of test cockroaches were carrying a higher bacterial load (1 × 104 and 1 × 105) as compared to control cockroaches. Multiple drug-resistant bacteria were isolated from test cockroaches. The diversity of drug-resistant bacterial species isolated from test cockroaches suggests their involvement in the transmission of drug-resistant bacteria. Various fungi and parasitic cysts of medical importance were also isolated from the test and control cockroaches, but the carriage rates were low. The findings suggest that cockroaches, in hospitals, can act as potential vectors of medically important bacteria/parasites/fungi.

Cats and dogs being treated at two veterinary clinics were investigated for gastrointestinal carriage of Clostridium difficile using selective solid and en-richment media. Thirty-two (39.5%) of 81 stool samples yielded C. difficile. There were significant differences in isolation rates between clinics, 61.0% of animals being positive at one clinic compared to 17.5% at the other (Chi-square, P < 0.005). Of 29 animals receiving antibiotics, 15 (52.0%) harboured C. difficile while 11 (23.9%) of 46 animals not receiving antibiotics were positive (Chi-square, P < 0.01). There was no difference in carriage rate between cats (38.1%) and dogs (40.0%). The environment at both veterinary clinics was surveyed for the presence of C. difficile. Fifteen of 20 sites at one clinic were positive compared to 6 of 14 sites at the other clinic. Both cytotoxigenic and noncytotoxigenic isolates of C. difficile were recovered from animals and environmental sites. These findings suggest that household pets may be a potentially significant reservoir of infection with C. difficile.

Duplicate neutralization tests were done on 401 avian and 101 human sera from island residents collected in the Coral Sea and on Australia's Great Barrier Reef against 19 known arboviruses. Antibodies to a potentially harmful flavivirus, Gadget's Gully virus, were equally present (4%) in both avian and human sera. Antibodies to another flavivirus, Murray Valley Encephalitis, and an ungrouped isolate, CSIRO 1499, were also present in both populations with non-significantly different incidences. Antibodies to Upolu, Johnston Atoll, Lake Clarendon, Taggert, Saumarez Reef and CSIRO 264 viruses were restricted to seabirds. Island residents with antibodies to Ross River and Barmah Forest viruses are thought to have been exposed to these viruses on the mainland as antibody to both viruses was absent among seabirds. These results indicate that consideration should be given to tick-associated arboviruses as potential public health hazards on islands where both seabird and human activities interact.