- Volume 24, Issue 4, 1987

A set of 25 Enterobacter cloacae typing phages was evaluated. Of 384 test strains, 93·8% were lysed by at least one phage; the mean number of reactions/strain was 7·3. Discrimination between strains was satisfactory within the most frequent O serotypes; 0·9 patterns/strain were found for strains of serotypes O3 and O8. Overall, 325 patterns were found. Phage patterns were completely reproducible when duplicates of strains were typed on the same day, but only 40% reproducible when repeated after 18 months. The combination of O serology and phage typing discriminated well between hospital isolates.

Immunisation of golden hamsters with plasma-membrane-associated antigens of a virulent subline of axenic Entamoeba histolytica strain NIH 200 V, entrapped in multilamellar phosphatidylcholine liposomes (MPL) or Freund’s complete adjuvant (FCA), afforded protection against intrahepatic challenge with axenic amoebic trophozoites of the same strain. Amoebic liver abscess developed in 86% and 80% of the animals that received empty liposomes or buffer emulsified in FCA but in none of the animals that received plasma-membrane-antigen vaccines. All the immunised animals had significantly higher levels (p < 0·001) of antibodies to plasma-membrane components and significantly higher levels (p < 0·001) of cellular sensitisation. Antibody-dependent macrophage-mediated cytotoxicity against trophozoites was also found to be significantly greater (p < 0·001) in immunised animals. Liposome-entrapped antigens stimulated the immune system of the host as well as, or better than, antigens administered with FCA.

The serotypes of strains of Campylobacter jejuni isolated from patients attending the International Center for Diarrhoeal Disease Research, Bangladesh (ICDDR, B), Dhaka, and from animals were ascertained by Penner and Hennessy’s (1980) serotyping scheme. Of 102 isolates from man, 74% were typable and serotypes 53, 15 and 22 predominated. Of 26 isolates from animals, 65% were typable and serotypes 15 and 53 occurred frequently. The diarrhoeal illnesses associated with different serotypes were similar. In one-third of cases other enteropathogens were present. C. jejuni was isolated from 7% of patients tested in a 4% sampling system during 1983, and the prevalent serotypes appeared in most months. It is concluded that C. jejuni is a common enteropathogen in Bangladesh, that a few serotypes predominate among isolates from both man and animals, and that serotype does not determine clinical symptoms.

Crude endotoxin preparations from Haemophilus actinomycetemcomitans and Bacteroides gingivalis showed activity in the two principal bio-assays for interleukin 1—the lymphocyte activating factor assay and stimulation of chondrocyte collagenase synthesis. Lipopolysaccharides purified from the crude endotoxins had reduced activity in the chondrocyte collagenase assay. The activity of the endotoxins may be due to synergic interaction between their lipopolysaccharides and other, as yet unidentified, bacterial components.

After exposure to cytotoxin or toxin A of Pseudomonas aeruginosa, the ultrastructure of resting and phagocytosing human polymorphonuclear leukocytes (PMNL) and of cells of P. aeruginosa strain 1348A was studied by transmission (TEM) and scanning (SEM) electronmicroscopy, and by light microscopy (LM) after histochemical staining of cytoplasmic granules. Cytotoxin caused marked clumping and destruction of PMNL, pyknotic nuclear changes with bleb formation, and release of cytoplasmic granules; phagocytosis was markedly diminished. In contrast, after exposure to toxin A, PMNL phagocytosed actively, but their cytoplasmic pseudopodia were markedly irregular and their nuclei pyknotic. Colloidal-gold-labelled cytotoxin showed an affinity for the cytoplasmic membranes, nuclei and granules of PMNL. Cytotoxin had no apparent effect on cells of P. aeruginosa strain 1348A but there was polar separation of the cytoplasmic membrane in bacteria exposed to toxin A. Cytotoxin and toxin A appear to be important in the pathogenesis of infections caused by P. aeruginosa.

Effects of four mucopolysaccharides and dextran sulphate on penicillin-induced lysis of Staphylococcus aureus FDA 209P were studied. Heparin and dextran sulphate inhibited lysis, whereas hyaluronic acid enhanced it. Chondroitin sulphates A and C had no effect. Incubation of S. aureus suspended in 0·03 m phosphate buffer (pH 7·0) with dextran sulphate inhibited autolysis of the bacteria, whereas incubation with hyaluronic acid enhanced autolysis. Both extracellular and cell-associated autolysin activities of S. aureus were suppressed by dextran sulphate and high concentrations of heparin. The addition of hyaluronic acid enhanced autolysin activity. The release of lipoteichoic acid (LTA), a modulator of autolysin activity, from penicillin-treated bacteria was inhibited by heparin and dextran sulphate. However, hyaluronic acid had no effect on release of LTA.

These results suggest that inhibition of penicillin-induced lysis of S. aureus by heparin results mainly from inhibition of LTA release while dextran sulphate inhibits both autolysin activity and LTA release. Hyaluronic acid appears to enhance penicillin-induced lysis through activation of the autolysins.

The association of Candida albicans with gastrointestinal (GI) mucosal surfaces was studied in vitro and in vivo. The caecal mucosal surfaces from antibiotic-treated and untreated control mice challenged orally with C. albicans revealed that large numbers of C. albicans were associated with the intestinal epithelium of antibiotic-treated mice but not with that of the control mice that possessed an indigenous wall-associated bacterial flora. Moreover, Candida cells only penetrated deep into the mucosa of animals in which the ecology of the intestinal microflora had been disrupted. In mice given antibiotics, C. albicans was associated with the mucosa of all areas of the GI tract; the caecal mucosa had the most associated Candida, whereas the stomach and small intestine had very few associated yeasts. Further examination of caecal mucosa from antibiotic-treated mice showed that C. albicans associated with the mucosa by at least five distinct mechanisms. These included: adhesion to epithelium, adhesion to mucus, co-adhesion to adherent fungi, co-adhesion to adherent bacteria, and entrapment in the mucous gel overlying the epithelium. The cell-surface hydrophobicity of C. albicans also was examined and found not to play a role in Candida adhesion to intestinal mucosa. The predominant association mechanisms appeared to be entrapment in the mucous gel, and adhesion to mucus and the epithelium. The ecological and pathological significance of co-adhesion by C. albicans to attached organisms is unclear but it may be important in the initiation of mucosal lesions.

Campylobacterpyloridis was cultured for maximal growth in liquid medium, and effects of exposure to various β-lactam and macrolide antibiotics, metronidazole, tripotassium dicitrato bismuthate (TDB) and cimetidine were monitored by transmission electronmicroscopy after periods of exposure up to 24 h. With amoxycillin and benzylpenicillin (0·12–1 mg/L) and cephalexin (2 mg/L) the normal bacilliform morphology was replaced by bulging and dumb-bell-like profiles showing cell-wall blebbing and vesiculation, and eventually by swollen forms with incomplete cell walls undergoing lysis. These changes developed progressively between 2h and 24 h and were accelerated at the higher antibiotic concentrations. Erythromycin and clindamycin caused central clearing, ribosomal coagulation and impaired cross-wall formation. There were no gross structural changes in the presence of metronidazole (4 mg/L), TDB (1000 and 2400 mg/L) or cimetidine (1000 and 2000 mg/L); but with TDB focal accumulation of particulate bismuth complex was detected under the cell wall, affecting nearly all organisms by 24 h. In parallel viability tests, metronidazole and TDB both showed bactericidal activity, but cimetidine did not. These findings support the clinical experience that favours combination therapy with bismuth plus an appropriate systemic antibiotic as the regimen of choice for effective clearance of the organisms in C. pyloridis-associated gastritis.

Clinical isolates of Neisseria gonorrhoeae harbouring resistance (R) plasmids of mol. wts 4·4 × 106 (Asian) or 3·2 × 106 (African) were grown in prolonged glucose-limited continuous culture to determine the segregation efficiency of each type of plasmid and their expression of penicillinase activity in the absence of antibiotic selective pressure. One strain contained the African plasmid and cryptic and conjugative plasmids, which were all retained after 96 generations in the chemostat. By contrast, the other strain lost all plasmids after 100 generations. Both strains showed increased sensitivity to a range of antibiotics, particularly to the penicillins. Loss of penicillinase activity as minimal inhibitory concentration decreased was confirmed for both strains by assaying the enzyme spectrophotometrically. Activity decreased with the number of generations and none was detectable at the time of complete plasmid loss. This decrease was apparently due to individual bacteria ceasing to produce enzyme rather than a gradual decline in production by the whole population. The sensitivities to a broad range of antibiotics also generally increased during glucose-limited growth, but one strain became more resistant to clindamycin and the other to tetracycline.

Strains of Escherichia coli isolated from documented cases of disease in pigs and belonging to a wide range of pathogenic serotypes were tested for their ability to produce a heat labile verotoxin (VT). The strains isolated from oedema disease all produced VT, indicating that the cytotoxin detected by the vero-cell assay was identical to “oedema disease principle”. Strains belonging to the serotypes associated with enterotoxic diarrhoea were VT–. Not all the strains belonging to the recognised oedema disease serotypes (O141:K85, O139:K82 and O138:K81) produced VT, but the VT– strains were not associated with outbreaks of clinical disease.

The behaviour of strains of Klebsiella aerogenes of capsular serotype K21 and strains of Escherichia coli producing a structurally related polysaccharide (colanic acid) was analysed by phagocytic and serum-killing assays. The cell-surface characteristics of these strains and of non-capsulate strains derived from them were also investigated by partitioning experiments in aqueous two-polymer phase systems. The possession of K21-type capsule by K. aerogenes or colanic-acid polysaccharide by E. coli conferred a strong negative charge on capsulate bacteria. Negatively charged bacteria of E. coli producing colanic-acid capsules, however, like non-capsulate K. aerogenes, were susceptible to uptake by polymorphonuclear leukocytes. In contrast, K21 polysaccharide conferred on klebsiellae considerable resistance to phagocytic uptake. The finding that ingested non-capsulate derivative strains of K. aerogenes were less rapidly degraded by phagocytes than E. coli strains suggested that other components of the cell surface of Klebsiella, notably lipopolysaccharide, may be involved in protection against phagocytic killing. The presence of colanic-acid capsules on E. coli conferred little resistance to the bactericidal activity of human serum or phagocytic uptake and did not protect against intracellular killing by polymorphonuclear leukocytes.