Infection control failures in a dental surgery--dilemmas in incident management.

Three separate incidents involving failure of decontamination of dental instruments were reported to our Unit in less than one year. We describe the risk assessment we undertook for the likelihood of detecting transmission of a blood borne virus infection. Even where 4000 patients attended the same dentist for seven years, there was no certainty of detecting even one person infected by the decontamination failure, while several people who had acquired infection by other routes would be identified. We conclude that these findings suggest that notifying patients is not usually justified.

Characterisation of strains of Aeromonas spp. by phenotype and whole-cell protein fingerprint.

Sixty-eight isolates of Aeromonas spp. were examined biochemically and their cell proteins were analysed by silver-stained SDS-PAGE. Protein fingerprints did not correlate with phenotype. However, consideration of both phenotype and fingerprint showed clustering of epidemiologically related isolates. There was also evidence that similar strains could be found in infected people and water or other environmental samples.

Whole-cell protein electrophoresis for typing Mycobacterium tuberculosis.

A method of discriminating between strains of Mycobacterium tuberculosis by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins combined with a sensitive silver stain is described. Thirty-five isolates of M. tuberculosis and five isolates from other species of Mycobacterium were examined, including serial isolates from the same patients and isolates from a small cluster of hospital cases. Different species of Mycobacterium were clearly distinguished, and within the species M. tuberculosis, different fingerprints were obtained, allowing discrimination of some strains from different patients. The reproducibility and discrimination of the technique are discussed.

Enterotoxic effects of Aeromonas sobria haemolysin in a rat jejunal perfusion system identified by specific neutralization with a monoclonal antibody.

Investigations into the pathogenesis of Aeromonas diarrhoea have demonstrated that several different cell-free products of motile aeromonads show enterotoxic activity in suckling mouse, rat, and rabbit assay systems. The relative contributions made by separate cytotoxic and cytotonic activities in the mixture produced by in vitro culture remains unresolved. Using a modified rat jejunal perfusion assay, we have studied the effects of A. sobria culture filtrates containing defined levels of haemolytic and cytotoxic activity and immunoreactivity for anti-cholera toxin. This material induced net water, potassium, and sodium loss with a rapid onset (less than 5 min) that was readily differentiated from the effects of purified cholera toxin (greater than 15 min). In filtrates containing up to 128 haemolytic and cytotoxic units of activity, the enterotoxic activity was neutralized by an anti-haemolysin/cytotoxin monoclonal antibody. No specific histological changes could be found in preparations perfused with enterotoxic material for up to 65 min. These findings indicate that the cytotoxic/haemolytic component of A. sobria culture filtrate is the dominant enterotoxic activity.

Direct selection of monoclonal antibodies neutralising the cytotoxic activity of Aeromonas sobria.

Monoclonal antibodies directed against the cytotoxic activity of Aeromonas sobria were raised by immunising mice with a culture supernatant concentrated by ammonium sulphate precipitation. Neutralising antibodies were specifically selected for by exposing hybridomas to cytotoxic levels of the immunising preparation. Cultures free from cytopathic effects after three hours were selected for further investigation. Ten cytotoxin resistant hybridomas were isolated but only two of these produced detectable neutralising activity in Vero and rabbit red blood cell assays. Different polypeptide binding patterns were observed for the neutralising antibodies compared with the other antibodies in immunoblotting studies. One of the neutralising antibodies was shown to act at an early stage in the development of cytotoxicity, probably by inhibiting binding.

Effects of incorporating ampicillin, bile salts and carbohydrates in media on the recognition and selection of Aeromonas spp. from faeces.

The effects of incorporating ampicillin, some bile salts and sugars into media on the primary recognition and selection of aeromonads from faeces were examined. Most (88%) of the 101 Aeromonas strains examined had an ampicillin MIC of greater than or equal to 40 mg/L, and would be isolated on blood agar containing ampicillin 30 mg/L. The strains with an ampicillin MIC of less than 40 mg/L were all of human origin and predominantly A. caviae. Although ampicillin at 10, 20 or 30 mg/L in blood agar inhibited faecal bacteria, the ability to detect Aeromonas strains with a high ampicillin MIC was less when the lower concentrations of ampicillin were used, without any improvement in the isolation of those strains with a low ampicillin MIC. Thirty-seven strains were tested for sensitivity to several different bile salts and found to be at least as resistant to them as Escherichia coli NCTC 10418. Bile salt sensitivity was not related to the species or source of a strain. There were minor differences in sensitivity to bile salts between some strains which related to whether strains had been isolated originally in the presence of bile salt or not. The effects of the presence of E. coli, Klebsiella spp. and Enterococcus faecalis on the growth of Aeromonas strains in mixed culture on media with and without carbohydrate were examined. The colony size of some Aeromonas strains was reduced in mixed culture but colony counts were not affected with any Aeromonas strains.(ABSTRACT TRUNCATED AT 250 WORDS)

Typing of Aeromonas species by protein fingerprinting: comparison of radiolabelling and silver staining for visualising proteins.

A method for typing Aeromonas species by silver staining of total soluble proteins separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis is described. There was good agreement with the results obtained by autoradiography of whole-cell proteins for isolates examined by both methods.

Haemagglutinating activity of Aeromonas spp. from different sources; attempted use as a typing system.

The haemagglutinating ability of 141 isolates of Aeromonas spp. for human, horse and guinea-pig erythrocytes was examined. Although the majority of isolates (136/141) agglutinated human group O erythrocytes, all the eight possible patterns of agglutination were observed. Haemagglutination of human group O erythrocytes, but not horse or guinea-pig, was associated with the ability to agglutinate yeast cells (Saccharomyces) and with aggregation in a low concentration of ammonium sulphate. Haemagglutinating ability was further characterized by reactions in the presence of mannose, galactose or fucose. All the possible patterns of inhibition with individual sugars were observed, but haemagglutination of human group O erythrocytes not inhibited by mannose, galactose or fucose was more common among isolates from patients with diarrhoea, and isolates producing a Vero cell cytotoxin than would be expected by chance. This might represent a virulence factor. When used as a typing system haemagglutination patterns did not correspond with the clustering of isolates expected on epidemiological grounds. Repeated subculturing resulted in a loss of agglutinating ability, an increase in the number of sugars inhibiting haemagglutination and an increase in the salt concentration required for aggregation. Haemagglutination did not seem to be useful as a means of typing aeromonads.

Epidemiology of Aeromonas species in a hospital.

The occurrence of Aeromonas spp. in a hospital water supply, in patients and in the local community was investigated. In healthy persons outside the hospital the isolation rate was 3.6% and among hospital patients it was 6%. Seven per cent of water samples yielded Aeromonas strains. Isolates were typed by sodium dodecyl sulphate polyacrylamide gel electrophoresis of sulphur-35 methionine labelled proteins of Aeromonas isolates. No relationship between water and human isolates could be established, even when a strain of A. hydrophila producing Vero cell cytotoxin contaminated enteral feeds given to patients in the intensive care unit.

Typing of Aeromonas species by polyacrylamide-gel electrophoresis of radiolabelled cell proteins.

A method for typing species of Aeromonas by 35S-labelling and sodium-dodecyl sulphate polyacrylamide-gel electrophoresis (SDS-PAGE) of total soluble proteins is described. Radiolabelled-protein profiles from different strains were compared by use of the Dice coefficient. Typability was 100% and discrimination of pairs of differing strains was satisfactory with acceptable reproducibility. The results also yielded information on subgroups of phenotypic species.

Isolation of Plesiomonas shigelloides from nutrient broth with brilliant green: its use in screening stool samples from an African population.

The suitability and selectivity of nutrient broth with bile salts and varying concentrations of brilliant green as an enrichment medium was tested for the isolation of Plesiomonas shigelloides from faeces. The broth was most effective at a concentration of 0.001 g/l brilliant green, using faeces deliberately contaminated with Plesiomonas. Two hundred and nineteen faecal samples from Nigeria were then examined for the presence of Plesiomonas and other bacterial pathogens. One yielded Plesiomonas following enrichment in bile salts brilliant green broth (BBG broth) whereas no samples were positive on direct plating.

Toxin production by Aeromonas spp. from different sources.

One hundred and eleven isolates of Aeromonas from water and from human sources were identified to species level and tested for the production of cytotoxin. These results were correlated with the source of each isolate and, for those from human faeces, with the clinical history of diarrhoea. A. caviae predominated in water, comprising 16 of 32 isolates; only one isolate from water was A. sobria. In human faecal samples 21 of 76 isolates were A. sobria; this was a significant difference. Cytotoxin producing strains were significantly more common in patients with no known cause for their gut symptoms. It is concluded that gastro-enteritis caused by Aeromonas is related to species and to production of cytotoxin.

Relationship of toxin production to species in the genus Aeromonas.

Ninety-five strains of Aeromonas were divided into three species--A. sobria, A. hydrophila and A. caviae--on the basis of results in 13 biochemical tests. The minimum number of tests necessary to distinguish these species was determined. Culture filtrate of the strains were tested for cytotoxin and cytotonin, haemolysin and protease. One filtrate with high-titre cytotoxin, haemolysin and protease activities was subjected to gel filtration on Sephadex G75 and isoelectric focussing. Of the five cell lines tested, Vero cells were most sensitive to the cytotoxin; no reproducible cytotonic effects were observed. The haemolysin effect appeared to be equivalent to cytotoxin. At least two distinct protease activities were found that might be responsible for the cytotonic effects described. Cytotoxin production was species related; it was present in A. sobria and A. hydrophila but not in A. caviae.

The colonization of patients in an intensive treatment unit with gram-negative flora: the significance of the oral route.

An extensive survey of patients and the environment in a newly refurbished intensive care unit showed that the principle species on patients in sites other than the rectum were Pseudomonas aeruginosa, Klebsiella pneumoniae, Serratia marcescens, Acinetobacter anitratus and Enterobacter cloacae. Multiple episodes of cross-infection were occurring with 10 different strains of these organisms. Three oral solutions (mouthwashes, 'Clinifeeds' and residual water from nasogastric aspiration apparatus) were heavily contaminated with coliforms including some epidemic strains and this corresponded with the finding that colonization with the above species usually occurred first in the mouth or respiratory tract. Attempts to eliminate contamination of the solutions reduced colonization and cross-infection by over 50%, but did not eradicate it. Two sinks without heat-traps on the drains possibly provided a long term reservoir of epidemic strains.

Aeromonas hydrophila in chlorinated water supplies.

Methods for the isolation and enumeration of Aeromonas hydrophila in water supplies are described. Examination of 286 chlorinated samples collected between July and October showed that 19% of otherwise uncontaminated waters contained Aerom. hydrophila, rising to 71% of those from which Escherichia coli was isolated. The isolation rate during January and February was only 7% for all samples. On the basis of these results, tap water is a likely source of the aeromonads found in human intestinal flora.

Methods for the isolation of Aeromonas hydrophila and Plesiomonas shigelloides from faeces.

Two solid selective media, xylose deoxycholate citrate agar (XDCA) and bile salts brilliant green agar (BBG) and an enrichment broth-alkaline peptone water, were evaluated for the isolation of Aeromonas hydrophila and Plesiomonas shigelloides. Alkaline peptone water and XDCA are useful for recovery of Aeromonas but not Plesiomonas, whereas BBG is satisfactory for both organisms.

Faecal carriage rate of Aeromonas hydrophila.

Over a four and a half month period, 1004 unselected routine faecal specimens from 815 patients were cultured for Aeromonas hydrophila. Forty-two specimens (4.2%) representing 38 patients were culture-positive. The study specimens also yielded Salmonella on 116 occasions, Shigella on seven, Campylobacter species on six and other bacterial pathogens on 17 occasions, respectively. Seven specimens had A hydrophila together with another bacterial pathogen. In only 19 of 38 patients (50%) was A hydrophila possibly associated with gastrointestinal symptoms. All the Aeromonas isolates were resistant to ampicillin but sensitive to gentamicin, piperacillin, mecillinam, chloramphenicol, ceftazidime and colistin.