- Volume 15, Issue 4, 1982

The inhibitory activity of seven strains of faecal streptococci against 34 strains of Clostridium difficile was examined in vitro after growth of the streptococci for 24 and 48 h. All strains of C. difficile were inhibited at 48 h but at 24 h the inhibition was variable. Streptococcus faecium, a group D streptococcus and an ungroupable streptococcus exhibited the most striking inhibitory activity. Lowering of pH of the medium occurred at the site of inhibition, but the pH change alone did not explain the inhibition of C. difficile. This antagonism may be related in vivo to the resistance to intestinal colonisation by C. difficile exhibited by the normal bowel flora, and in vitro to failures to isolate C. difficile from faecal specimens.

The use of eight rapid tests for the identification of 1307 strains of mycobacteria belonging to 18 species was evaluated. The standard niacin, nitrate-reductase and catalase tests were supplemented by new tests for the detection of β glucosidase, urease, penicillinase, trehalase and cephalosporinase. This combination of eight rapid tests was not able to replace more conventional procedures but in some cases was of value in discriminating between closely related species.

Contrary to current opinion, neutrophil leucocytes from patients with the inherited immunodeficiency syndrome chronic granulomatous disease (CGD) killed 80% of ingested Staphylococcus aureus. Bacterial killing was not impaired by increasing the ratio of bacteria to cells from 1:1 to 10:1. The organisms that survived within the patients' cells did not themselves appear to constitute an unduly resistant subpopulation because they were killed when exposed to fresh cells, and no growth phase of a synchronous culture was found to be particularly resistant. The pH within the phagocytic vacuoles of CGD neutrophils and monocytes is abnormally low and methylamine, which has been shown to normalise this vacuolar pH, improved killing. Clumped bacteria appeared to be more resistant to killing than dispersed ones, suggesting that organisms near the centre of a clump might be protected from the toxicity of the compromised killing systems in cells of these patients.

Cell-bound opacity factor (OF) was extracted with sodium dodecyl sulphate (SDS) to yield stable extracts with titres of > 20 000. The mol.-wt distributions of extracellular and SDS-extracted OF, determined by ultrafiltration or chromatography on Sepharose 4B, suggested that the high mol. wt (1 × 106) of extracellular OF is due to aggregation, because cell-bound and extracellular OF in the presence of SDS had an average mol. wt of only 2 × 105.

At least four apparent multiple-molecular forms (mol. wt 7.4-12.0 × 104) of OF were detected by SDS polyacrylamide-gel electro-phoresis. It seemed more probable that these were due to aggregation rather than the existence of different stable conformations. To explain the molecular-size distribution, the subunit would have to be as small as 1 × 104 but this was supported by the finding that OF can be detected after passing through a dialysis membrane provided that its “substrate”, α1-lipoprotein, is present on the other side. This raises the possibility that OF is associated with a carrier molecule.

The isoelectric-focusing profiles of OF were complex and differed markedly with the method used to prepare OF. Extracellular OF had a simple profile with an isoelectric point of 4.0, whereas Triton-extracted OF was the most complex and formed three peaks, the position of which varied depending on whether the detergent was present or absent during focusing runs.

The comparative pathogenicity of different species of Actinomyces was studied in a susceptible weanling-mouse model. After the intraperitoneal injection of strains of Actinomyces israeli, A. naeslundi, A. viscosus and Arachnia propionica, numerous abscesses developed in the intestine, mesentery, liver, and at the site of injection. Lesions were not produced by A. odontolyticus. A. naeslundi and A. viscosus produced acute lesions that resolved after a few weeks. Abscesses produced by rough strains of A. israeli and Arach. propionica persisted and led to a slowly progressive chronic infection. Viable organisms were always recovered from the lesions. Spread of the lesions by extension into other areas, including the thoracic cavity, led to the death of the animal after approximately 1 year. This study demonstrated a clear difference in the pattern of infection produced by the different species of Actinomyces as well as Arach. propionica.

Resistance to 11 antimicrobial drugs was assessed in 532 clinically significant strains of Staphylococcus epidermidis received in the years 1978, 1979 and 1980 and compared with that of strains collected in 1976 and 1977 for an international study. Only 14% of strains were sensitive to all the drugs tested. Resistance to gentamicin increased significantly from 7% to 33% during the study period. The degree of association between pairs of drugs was assessed. There was strong association between resistance to methicillin, aminoglycosides and macrolide antibiotics and only weak association between resistance to novobiocin, tetracycline, chloramphenicol and penicillin. Patterns of resistance were complex and may be useful as an accessory typing system.

The genus Corynebacterium, defined by the presence of mesodiaminopimelic acid, arabinose and corynomycolic acids in the cell wall, contains a very diverse group of organisms according to the results of numerical analysis of protein patterns; 13 groups were distinguished in this study. Strains isolated from axillary skin and hair included representatives of several groups but many strains were placed in groups 1 and 6A. Neither of these groups contained any reference strains and may constitute hitherto undescribed species. The reference strains of C. diphtheriae formed a coherent group not closely related to any non-pathogenic Corynebacterium species.

Leptospira interrogans serovars pomona and hardjo were adapted to grow in a chemically-defined, protein-free (PF) medium. Formolised monovalent vaccines of serovars pomona and hardjo and a bivalent mixture of the two were prepared from PF cultures. Live PF cultures and the vaccine preparations retained their agglutinating antigens and their immunogenicity when tested in rabbits and guineapigs. The vaccines were not pyrogenic and dermal reactions were slight.

Clostridium perfringens strains resistant to metronidazole and tinidazole were isolated from the sensitive parent strain CM288 after mutagenesis with N-methyl-N′-nitro-N-nitrosoguanidine. Strain CM288 was already resistant to rifampicin and nalidixic acid; these genetic markers helped to confirm the identity of mutants. All mutants showed similar characteristics: they grew more slowly than the parent strain and failed to reach the same maximum turbidity; uptake of metronidazole and tinidazole from culture fluids was slow and end products of glucose metabolism were different from those of the parent. Pyruvate dehydrogenase activity was not detected in broken cell preparations of the mutant strains although this enzyme was readily detected in the parent strain. Changes in end products of glucose metabolism were consistent with the absence of pyruvate dehydrogenase activity because pyruvate was accumulated during growth and lactate levels were higher whereas acetate, CO2 and ethanol levels were diminished.

The effects of dietary carbohydrates on the adherence of Candida albicans to HeLa epithelial monolayers and buccal epithelial cells were compared by two assay systems. Candida preincubated in 0·5m, glucose, sucrose, galactose, xylitol or maltose medium produced a significant enhancement in adhesion to both types of epithelial cells. Maltose was the most effective sugar and glucose the least effective in promoting adhesion, while lactose had no significant effect. A clinical isolate of C. albicans demonstrated a greater overall enhancement in adhesion from preincubation with glucose, sucrose and maltose, when compared with a reference laboratory strain of Candida. These results imply that exogenous or endogenous carbon sources may affect the oral and vaginal carriage of C. albicans, by modifying their adhesive properties.

Twenty-eight pre-term babies of low birth weight were monitored for developing microflora in throat, stomach and faeces during the first 3 weeks of life. The flora at all levels of the gastrointestinal tract differed from that of healthy breast-fed and artificially fed full-term babies. Colonisation of throat and stomach was delayed beyond 4 days of life in 87% and 60% of babies respectively. Only 10% of babies had “normal” oral flora throughout the period of study. Flora of the stomach was sparse, and resembled faecal flora. Faecal flora was established more rapidly than throat or stomach flora, and 70% of babies were colonised during the first 4 days of life. Initially Bacteroides spp. were predominant (57% babies), but Escherichia coli and other aerobic gram-negative bacilli gradually increased in frequency. Colonisation by gram-positive bacteria was slow. Clostridium spp. were present in only 10% of babies during the first 4 days of life. Most strains were transient. Colonisation with C. butyricum (30%), C. perfringens (35%) and C. difficile (25%) was maximum after the first 2 weeks of life. Lactic-acid-producing bacteria usually appeared late in the third week of life. Parenteral feeding immediately after birth was associated with delayed colonisation by a restricted number of species. Parenteral antibiotics (penicillin or gentamicin or both) restricted colonisation with normal oral flora, the lactic-acid-producing bacteria and penicillin-sensitive clostridia, but had little effect on E. coli even when the colonising strain was sensitive to the aminoglycoside in the regimen. Systemic spread of bacteria via the blood stream was not detected in any babies.

The pattern of colonisation of the enteric tract in pre-term infants in the special-care nursery studied, differs from that of healthy full-term babies; this merits consideration when the results of bacteriological tests on this vulnerable group of infants are being interpreted.

The effects of the thermolabile (LT) and thermostable (ST) enterotoxins and the Vero-cell (VT) toxin of Escherichia coli were studied in continuous cell lines in tissue culture. The LT enterotoxin induced morphological changes in mouse adrenal (Y-1), Chinese hamster ovary (CHO-K1) and African green-monkey kidney (Vero) cells. VT toxin produced cytotoxic effects in adult Rhesus-monkey kidney (LLC-MK2), human embryonic foreskin (HFS) and Vero cells. The ST enterotoxin had no effect on any of the cell lines.

The detection of gonococcal antigens by an indirect sandwich ELISA system is described. The feasibility of using rabbit antiserum raised against whole cells of Neisseria gonorrhoeae strain 9 to detect gonococcal lipopolysaccharide, whole cells, and outer membrane (OM) protein was investigated. OM protein was found to be the main antigen detectable with the antiserum in a direct ELISA system. A positive result in the indirect assay could be obtained with a minimum of 46 to 92 ng of gonococcal OM protein or with 6·6 × 103 cfu of N. gonorrhoeae. The sensitivity of the assay was found to be approximately eightfold lower with OM complex from a strain of N. meningitidis serogroup B for which the minimum amount of OM protein detected was 375 ng. Negative results were obtained with OM complex from Streptococcus agalactiae, Bacteroides bivius and Escherichia coli. The assay seems to be highly specific for gonococcal antigens. The sensitivity of the assay and its specificity commend it for further evaluation in the detection of gonococcal antigens in clinical specimens.

One hundred and thirteen strains of Morganella, Proteus and Providencia, grown in different cultural conditions, were examined for their ability to produce haemagglutinins (HAs). Three main kinds of HA (MS, MR/K and MR/P) were detected, and 89% of the 112 HA+ strains were capable of producing two or three of the different HAs in the same or different cultures. The properties of the three HAs were partly defined and the difficulties of identifying their separate HA activities when present together are discussed. Electronmicroscopic examination of bacteria from HA+ cultures showed at least six distinct fimbrial types, the properties of which are described. We tried, with limited success, to correlate the presence of the different HAs with that of the different fimbrial types. The significance of our findings is reviewed in the light of recent taxonomic changes for this group of enterobacteria. The distribution of HAs and fimbriae in the species of Morganella, Proteus and Providencia is more complex than that so far described for other genera of Enterobacteriaceae.