Epidemiology of an outbreak of Clostridium difficile-associated-diarrhoea. Application of a typing system.
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Biomedical subjects
Publications and source records attributed to S Tabaqchali.
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An outbreak of septicaemia with Pseudomonas aeruginosa amongst adult men with haematological malignancy involved eight patients on the same ward during a period of 5 weeks. The strains isolated from blood cultures from seven patients were indistinguishable by conventional typing methods. Thymol mouthwash which had been made up and distributed in communal jugs was found to be contaminated with the epidemic strain and was the likely source for this outbreak. A high rate of gastrointestinal colonization with the epidemic strain was found in the patients receiving the contaminated mouthwash. Only those patients with prolonged severe leucopenia developed septicaemia. Communal medications are an unnecessary hazard, particularly in oncology wards.
The serological response to Aspergillus fumigatus was investigated in patients with pulmonary aspergillosis using the immunoblot technique. Antibody was detected against nine components of the fungus, ranging in molecular weight from 88 000 to 33 000 daltons. Antibody to a 40 000 dalton component was present in most patients (13 of 16). In surviving patients it was high (two with aspergilloma) or low but persistent (two with invasive aspergillosis). It was absent or fell in the 12 patients who died of invasive aspergillosis. Antibody to this 40 000 dalton antigen was not present in 10 sera from healthy controls. This antigen may form the basis of a more sensitive and specific serodiagnostic test for systemic aspergillosis.
Western blotting was applied to the analysis of Candida albicans in the yeast and mycelial phases in an attempt to recognise mycelial specific antigens which might be of serodiagnostic value. The antisera were prepared in rabbits by immunising them with pressates of C albicans type A NCTC 3153 in the yeast phase or the mycelial phase. These were blotted against C albicans in the yeast and mycelial phases and the yeast phase of C parapsilosis, C krusei, C tropicalis, and Torulopsis glabrata. Cross reactivity was greatest against C parapsilosis. One yeast specific mannoprotein was identified with a molecular weight of 49 000. No mycelial specific antigens could be identified.
Volatile fatty acids produced in Robertson's cooked meat medium by a range of clinically relevant anaerobes were compared by gas liquid chromatography with those produced in blood agar. The same volatile fatty acid profiles were obtained in both media, although the concentration of acids was lower in blood agar. We conclude that detection of volatile fatty acids from a pure culture of an organism on solid medium is practicable and offers advantages over the conventional technique.
To investigate the heterogeneity in antibody response to the various antigenic determinants of Candida albicans in patients with disseminated candidosis 201 serial serum samples from 45 patients with proven systemic candidosis were examined by immunoblotting (Western blotting) for antibodies to C albicans type A NCTC 3153. 40 patients had detectable antibody. 26 different antigen bands, ranging from 104 kD to 23 kD, were identified. Despite wide variation in the antigens recognised by different patients, the antibody responses fell into six patterns (A to F). 13 patients infected during an outbreak of systemic candidosis gave the same response (A). Of a further 7 patients with this response, 2 had isolates indistinguishable from the outbreak strain. Group B responses were associated with leukaemias and lymphomas. Groups C-F responses were seen in 9 patients. Production of antibody to a 47 kD antigen, common to all six groups, occurred in all those who recovered from the infection and may therefore be of prognostic significance. Whether this antigen could be used as the basis of a vaccine remains to be determined.
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Epidemiological studies of Clostridium difficile diarrhoeal disease have been hindered by the lack of a typing scheme for this organism. A typing method based on the incorporation of sulphur-35-labelled methionine into cellular proteins and their separation by sodium dodecylsulphate/polyacrylamide gel electrophoresis showed clear pattern differences between strains, and nine distinct groups within the C difficile species were established. 98% of 250 clinical strains derived from four hospitals were typable. Group X was the commonest group and was associated with outbreaks of pseudomembranous colitis and antibiotic-associated colitis in two hospitals. Groups A-D were isolated predominantly from mothers and newborn infants. In outbreaks of antibiotic-associated colitis in oncology and orthopaedic wards the same strains, group X and group E, respectively, were isolated from patients and their environment, providing strong evidence of cross-infection between patients and of hospital acquisition of C difficile.
Specimens of mucosal tissue of the small and large bowel obtained either by a colonoscope or during surgical operations were investigated quantitatively for their bacterial flora. A detailed prescription of the methodology using an anaerobic chamber is given. Most of the mucosal specimens of the small intestine as well as all the specimens of the large intestine contained bacteria (about 10(4) germs/g). Predominantly, Bacteroides, gram-positive rods, and cocci were isolated. In contrast to the gut lumen flora anaerobic bacteria of the mucosal flora did not outnumber the aerobes. In patients with inflammatory bowel disease or with carcinoma of the colon the mucosal flora showed no demonstrable alterations even after undergoing a sulphonamide therapy.
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The determinant responsible for the ability of Bacteroides spp. to inhibit polymorph phagocytic killing of aerobic organisms has not yet been identified. Therefore, the roles of lipopolysaccharide and capsular polysaccharide of B. fragilis were investigated. Serum-resistant and serum-sensitive strains of Proteus mirabilis were used to indicate inhibition of phagocytic killing and serum killing of aerobes. Whole organisms of B. fragilis, purified lipopolysaccharide and capsular polysaccharide were added to an in-vitro phagocytosis system. Results showed that greater than 10(7) bacteroides/ml inhibited both serum and phagocytic killing. Concentrations below 10(7)/ml had little effect on either process. Purified capsular polysaccharide (10 or 100 micrograms/ml), either alone in the system or in combination with sub-inhibitory concentrations of B. fragilis also markedly inhibited serum and phagocytic killing. Lipopolysaccharide (9 micrograms/ml) appeared relatively inert. B. ovatus, reputedly non-capsulated, produced identical results to those obtained with B. fragilis, but an encapsulated strain of Streptococcus pneumoniae did not inhibit serum or phagocytic killing.
The relationship between vaginal carriage and subsequent neonatal acquisition of Clostridium difficile was investigated. Vaginal carriage of C. difficile was detected in 11% of women attending the Department of Genital Medicine Clinic. C. difficile was isolated from the vagina in 18% of 50 mothers before delivery, and 8% after delivery; 62% of their babies had positive faecal cultures. Eight of nine of the babies whose mothers had positive cultures before delivery became colonised with C. difficile, while 23 of 41 babies whose mothers had negative cultures became colonised. This suggests that both the vagina and the environment may act as sources of neonatal acquisition of C. difficile. Broth enrichment culture proved a more sensitive method for isolating C. difficile from the vagina than direct plate culture and should be used in such investigations.
Samples from the posterior vaginal fornix of 102 women with various clinical conditions were analysed by a quantitative method. Aerobes were isolated from all but one of the specimens at a mean concentration of 7.2 log10 cfu/g and anaerobes from 92 specimens at a mean concentration of 8.1 log10 cfu/g. In most clinical conditions and in a control group of asymptomatic women, anaerobes outnumbered aerobes by about ten to one (one log10 unit). The most common organisms were aerobic and anaerobic lactobacilli, coryneforms, Staphylococcus epidermidis, Bacteroides spp. and anaerobic gram-positive cocci. Lactobacilli did not appear to confer any protective effect by excluding the presence of other organisms such as Gardnerella vaginalis or anaerobes. The isolation of anaerobic organisms from the vagina cannot be regarded as being of pathogenic significance without other supporting evidence.
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Pyruvate dehydrogenase activity was present and comparable (giving a colour change of 202-673 od units/min/g protein in our assay system) in two clinical strains of Bacteroides fragilis exhibiting reduced sensitivity to metronidazole (MIC 2-8 mg/l at 48 h) in a resistant strain (Ingham's) of Bact. fragilis (MIC 64/mg/l) and in two sensitive strains of Bact. fragilis (MIC 0.5 mg/l). Pyruvate dehydrogenase activity was not detected in a capnocytophaga strain (sensitive to 50 micrograms disc of metronidazole), in Escherichia coli or in 'Bact. fragilis' strain (Britz) AM24s (MIC greater than 64 mg/l). Further testing of AM24s strain indicated that it was not Bact. fragilis but a facultatively anaerobic Gram-negative rod. Pyruvate dehydrogenase activity therefore, is similar in sensitive and resistant strains of Bact. fragilis and is unlikely to be related to the mechanism of resistance to metronidazole.
Rectal swabs from 122 patients and 497 environmental swabs from several wards were examined for the presence of Clostridium difficile in order to assess the role of the environment in the spread of this organism. Clostridium difficile was isolated from 6/27 (22.2%) oncology patients and from 8/163 (4.9%) environmental specimens obtained from the oncology unit. Items found positive for C difficile were those subjected to faecal contamination such as commode chairs, bed pans, dust pans, discard bins, the sluice and a disposable bed pan machine. Fourteen of 51 (27.4%) asymptomatic neonates yielded mostly toxigenic C difficile in their stools during their first week of life. Five of 156 (3.2%) specimens taken from inanimate objects in the environment of the neonatal units were positive for C difficile. The organism was also isolated from the hands of a nurse. Similar antibiogram patterns were demonstrated in the strains obtained from the patients and their environment indicating the possible occurrence of cross infection. These results indicate that environmental contamination is important in the spread of C difficile in hospitalised patients and the implementation of isolation procedures may limit that spread.