- Volume 21, Issue 3, 1986

The slime glycolipoproteins (GLPs) extracted from Pseudomonas aeruginosa strain C2 and its laboratory-induced gentamicin-resistant variant were analysed for gross chemical composition. The GLP of the wild-type strain contained significantly greater amounts of neutral sugars, uronic acid and thiobarbituric-reactive material (p<.001) than the GLP of the gentamicin-resistant variant. Also significantly higher (p<.01) was the amino-sugar content of the GLP from the wild-type strain. Paper chromatographic analyses of the hydrolysates of the GLPs revealed that two neutral sugars, rhamnose andmannose, were absent from the GLP of the resistant variant. The GLP of strain C2 contained significantly less protein than the GLP of the gentamicin-resistant variant.

Inducibility of β-lactamase activity by cefoxitin was examined in 626 gramnegative clinical isolates selected for amoxycillin and cephalothin resistance. The results indicated that precise identification and cefoxitin sensitivity or resistance could be used to predict the inducibility of β-lactamase. Of 326 organisms from species capable of β-lactamase induction, inductionwas shown in 68% and was predictable from the cefoxitin-sensitivity and identification data. No induction of β-lactamase occurred in the remaining species. Acomparison of β-lactamase activities against cefotaxime, cefoperazone and latamoxef showed that inductionofenzyme activity against cefotaxime and cefoperazone occurred at similar rates. Induction of activity against latamoxef did not occur or was minimal with three bacterialspecies. The data show that of 119 strains of Enterobacteriaceae displaying inducible β-lactamase, 113 would have been reported as unequivocally sensitive to cefotaxime, 109 as sensitive to cefoperazone and 116 as sensitive to latamoxef if the disk-diffusion technique alone hadbeen used. The majority of Pseudomonas strains examined produced inducible enzyme and they were moreresistant to the three cephalosporins tested than were the Enterobacteriaceae.

The effect of saliva and serum on the adherence of five strains of Candida albicans and one each of C. tropicalis and C. glabrata to chlorhexidine-pretreated acrylic was measured in vitro. A four-fold dilution of saliva or serum significantly inactivated the fungicidal effect of chlorhexidine gluconate. Pretreatment of the acrylic with unstimulated mixed saliva for 30 min led to a reduced adherence for all the Candida strains tested, whilst a similar pretreatment with serum slightly increased adhesion. Moreover treatment of saliva- or serum-coated acrylic with chlorhexidine gluconate 2% reduced adherence by between 19% and 86%. The inhibition of yeast adherence by chlorhexidine persisted for up to 19 days after the exposure of the acrylic strips to the disinfectant.

Blood-culture broths showing macroscopic or radiometric evidence of growth of gram-negative bacilli were examined by a rapid automated bacterial identification system. A differential centrifugation technique was developed to prepare suitable inocula. The identification results obtained were confirmed by the API 20E method, with single colonies of the strains isolated 24 h later. Of 90 organisms tested, seven did not give the same identification by the two systems. With the rapid automated technique a presumptive identification of gram-negative bacilli can be made 24 h earlier than by more conventional methods.

Haemophilus strains isolated from children under the age of 11 months with conjunctivitis were characterised by biotype, sugar fermentation, plasmid pattern and outer-membrane-protein profiles. H. influenzae was the most common species identified and was separated into 14 groups based on sugar fermentation and biotype patterns and into more than 20 groups when plasmid and outer-membrane-protein profiles were included. Small (mol. wt< 10 × 106) plasmids were identified in 11 of 34 (32%) H. influenzae isolates, 1 of 2 H. haemolyticus and 4 of 6 (67%) H. parainfluenzae isolates. Examination of sugar-fermentation and plasmid patterns increased the ability to distinguish between strains isolated at different times from recurrent disease and may have general applications in the study of Haemophilus strains isolated from a single anatomical site.

Cross-reactivity in a delayed-type hypersensitivity (DH) response was studied in mice immunised with live Salmonella typhi, S. paratyphi A and S. paratyphi B. Extensive cross-reactions outside the serogroup limits were observed. The ability of DH cross-reacting and non-cross-reacting sonicates to generate activated macrophages was studied in mice immunised 3 months earlier with S. paratyphi B. Whereas DH cross-reacting S. poona sonicate generated activated macrophages the non-cross-reacting S. typhi sonicate did not. To determine whether infections due to diarrhoea-causing salmonellae generated cross-reactive cell-mediated immune responses against enteric fever-causing organisms, similar reverse experiments were performed in mice immunised with S. enteritidis. S. paratyphi A sonicate generated both effector responses, i.e., DH and activated macrophages.

The virulence (expressed as LD50 values) for mice of two mutant strains of Salmonella dublin, both containing TnA insertions in the resident plasmid, was reduced by 104— 105 when infection was by the oral or intravenous or intraperitoneal route. When the plasmid was lost from one of the mutants no further decrease in virulence was observed. Results also suggested that plasmid genes are not involved in the ability of S. dublin to cross the gut wall.

The role of Bacteroides fragilis in the pathogenesis of acute appendicitis was studied in 135 patients in four patient groups: normal (17); phlegmonous appendicitis (17); gangrenous appendicitis (75); and septic complications of appendicitis (26). Aerobic and anaerobic bacteria were isolated from all groups and members of the ‘B. fragilis group’ were the most common anaerobic isolates. The rate of isolation of B. fragilis was similar from normal and inflamed appendices but was significantly higher from those with septic complications (p<0-01). Antibodies against B. fragilis were demonstrated in patients of all groups and occurred with similar frequencies in patients with normal and inflamed appendices but at a significantly higher rate in those with septic complications (p<0 01). Whereas patients in this latter group showed IgM- antibody responses to B. fragilis only, those with acute appendicitis had IgM antibodies against a wide range of organisms of the 'B. fragilis group’ which suggests that B. fragilis does not play a significant role in acute appendicitis but may be a major cause of its septic complications.

Laboratory strains of Mycobacterium phlei, M. smegmatis, M. fortuitum, M. gordonae, M. kansasi, M. bovis, M. tuberculosis and M. intracellulare were adapted to grow in an anaerobic environment. Concomitant with the transition to anaerobic growth was loss of acid-fastness, loss or modification of colonial pigmentation, and loss of ability to grow on a malachite green-containing medium. The mycobacteria grown anaerobically produced acid from a greater range of carbohydrates than aerobically grown cultures, lost iron-uptake activity, and showed a reduction of urease, catalase and nitratase activity. Back adaption of mycobacteria from an anaerobic to an aerobic environment resulted in the acquisition of acid-fastness, pigmentation, and other characteristics used in the taxonomy of mycobacteria. These results suggest that mycobacterial cultures, if grown in an anaerobic environment, may be erroneously identified in clinical laboratories.

Antibody activity against Mycobacterium tuberculosis of sera from an area with a high prevalence of tuberculosis was measured by enzyme-linked immunosorbent assay (ELISA) with a plasma-membrane extract from M. tuberculosis strain H37RV. All sera from relapsed tuberculosis patients and 82.5% of sera from new untreated cases gave positive results. The seronegative group of tuberculosis patients gave positive results by direct microscopy and culture. No clear correlation between antibody and delayed hypersensitivity or extent of disease was observed. Chemotherapy was associated with a higher antibody response. Specificity of the test with healthy control subjects from the high prevalence area was 85%. Negative results were obtained with 145 sera from presumed healthy European subjects and with seven sera from BCG-vaccinated subjects.

Bordetella pertussis heat-labile toxin (HLT) was assayed by the haemorrhagic response produced by subcutaneous injection into weaned mice. Young mice, 3-5 weeks old of either sex, were highly responsive but they became resistant to HLT as they matured. Two anti-inflammatory agents, prednisolone and meclofenamate, inhibited the skin reactions in young mice. When given intraperitoneally, prednisolone was most inhibitory if it was injected just before, or at the same time as, HLT challenge. Prednisolone given 3 h after challenge, when the skin reactions had started to develop, did not significantly attentuate the final response. Both drugs were even more effective when mixed with the HLT challenge and injected subcutaneously. These findings are discussed in relation to the possible mechanism of action of HLT and to the reported beneficial effects of corticosteroids in the treatment of whooping cough.

A new enzyme-linked fluorescence assay (ELFA) suitablefor use with peroxidase-antibody conjugates is described.The substrate for the assay is p-hydroxyphenylacetic acid, the fluorescent product of which is stable and unaffected by light. The assay compared favourably with a standard ELISA for the quantitation of IgM antibodies to hepatitis B core antigen.

Ten antisera, prepared against nine selected strains of group-G streptococci and a strain of group-A streptococci of M-type 12, were used to serotype 102 isolates of group-G streptococci by means of precipitation reactions between the sera and hot-acid extracts of the streptococci.Fifty-six (54.9%) of the streptococci could be typed; eight serotypes were identified, of which type VIII was the most common (18.6%). The M12 and the R28 antigens, previously recorded in group-G streptococci, were not detected. The type antigens were trypsin sensitive and resembled the M antigens of group-A streptococci.