Tests of performance of anaerobic jars.

This paper briefly reviews methods of assessing the in-use performance of anaerobic jars and outlines a simple system combining a rapid test of catalytic activity with a biological indicator that can detect defects in the jars after incubation.

Recovery of anaerobic bacteria from small inocula: a model for blood culture studies.

The recovery of anaerobic, facultative anaerobic, and aerobic pathogens from very small inocula was studied under conditions that could be related to routine blood culture procedures. Results with Brain Heart Infusion broth were unsatisfactory. Freshly prepared Brewer thioglycollate medium gave apparently good results, but there are disadvantages when this medium is used for blood culture. Results with Difco Thiol broth were disappointing. A modification of Robertson's cooked-meat broth supplemented with Brain Heart Infusion gave good recovery and sustained viability with a wide range of test organisms including exacting strains. The findings raise points of practical importance.

The production of neuraminidase by food poisoning strains of Clostridium welchii (C. perfringens).

The production of neuraminidase by a classical strain of Clostridium welchii (C. perfringens) type A was studied. Good yields were produced in 5% Proteose Peptone-water medium (PPW5); the enzyme was essentially extracellular but some further neuraminidase could be released by ultrasonic disintegration of the cells. This also released N-acyl neuraminic acid-aldolase (NAN-aldolase) and the degree to which this interferes with the assay for neuraminidase was evaluated. Forty-one British reference food-poisoning strains of C. welchii type A were examined for extracellular neuraminidase production in PPW5. Twelve of 17 strains that produce so-called heat-sensitive spores were neuraminidase positive whereas 20 of 24 strains that are non-haemolytic and produce very heat-resistant sporeswere neuraminidase negative. Variation was found in the ability to produce neuraminidase among strains of a single Hobbs' serotype; four Hobbs' type-13 strains produced neuraminidase but a fifth did not. Disruption of the cells of a Hobbs' type-2 strain that did not produce any extracellular neuraminidase released NAN-aldolase but there was no evidence of cell-associated neuraminidase. British food-poisoning strains of C. welchii type A thus include some that are clearly neuraminidase positive and some that still cannot be shown to produce neuraminidase. There is no correlation between lack of neuraminidase production and the ability to cause food poisoning, although the majority of non-haemolytic heat-resistant strains do not produce neuraminidase. It remains possible that neuraminidase may play a part in C. welchii gas gangrene; it is suggested that the ability to define neuraminidase-negative strains may now be of value in investigating this possibility.

Detection of species-specific and cross-reactive cell-surface antigens of Bacteroides species by an indirect enzyme-linked immunosorbent assay.

EDTA-released outer-membrane antigen complexes were prepared from 172 laboratory and reference strains and 43 fresh clinical or faecal isolates representing 20 species or subspecies of Bacteroides, together with 13 species of other genera. These antigens were titrated against antisera to whole, live cells of 21 species or subspecies of Bacteroidaceae in an indirect enzyme-linked immunosorbent assay (ELISA). The presence of species-specific antigens was investigated and cross reactions between species were noted. Results showed that a high proportion of the species possess species-specific antigens with little significant cross reactivity. We believe that the ELISA system described here detects antigens that represent the whole cell surface of bacteroides organisms. We have exploited the sensitivity of the system and its quantitative potential to define approaches for those wishing to use serological approaches for either the identification of Bacteroides species or for the titration of serum antibodies in patients with a possible bacteroides infection.

Immunochemical fingerprinting of Clostridium difficile strains isolated from an outbreak of antibiotic-associated colitis and diarrhoea.

Twenty eight strains of Clostridium difficile , isolated from an outbreak of antibiotic-associated colitis and diarrhoea in an orthopaedic ward and from sporadic cases throughout Sweden, were sent to Edinburgh for immunochemical fingerprinting without information about their origin. EDTA extracts of the organisms were examined by crossed immunoelectrophoresis (CIE), polyacrylamide gel electrophoresis (PAGE) and electroblot transfer. Two patterns were revealed by CIE: group A (18 strains) and group B (10 strains). PAGE and electroblot transfer revealed one major group of 10 strains (group 1), six small groups of two or three strains and six strains which were unlike any other strain. The CIE group B and PAGE- electroblot group 1 were identical. Nine of the 10 strains in this group were from patients in the outbreak. These findings indicate that a single strain spread in the orthopaedic ward as a nosocomial infection and that this strain differed from most other strains investigated. The PAGE- electroblot technique should, therefore, greatly aid investigations into the epidemiology of C. difficile infections.

A scheme for the identification of clinical isolates of Gram-negative anaerobic bacilli by conventional bacteriological tests.

More than 1000 strains of gram-negative anaerobic bacilli, including reference strains, clinical isolates, and members of the normal flora of the mouth, lower gastro-intestinal tract and vagina of healthy human subjects, were studied by conventional bacteriological methods and by gas-liquid chromatographic analysis of metabolic products in a series of investigations. A short combined set of tests with particular discriminant value was selected, and a scheme for the identification of the species and subspecies encountered in the diagnostic bacteriological laboratory was based upon our composite results. The tests are: antibiotic-disk resistance tests with neomycin 1000 micrograms, kanamycin 1000 micrograms, penicillin 2 units and rifampicin 15 micrograms per disk; tolerance tests with sodium taurocholate, Victoria blue 4R and gentian violet; and tests for pigment production, indole production, aesculin hydrolysis and the fermentation of glucose, lactose, sucrose, rhamnose, trehalose, mannitol and xylose. Gram-negative anaerobic bacilli are divided into four groups: (1) the fragilis group with nine species, which include the five subgroups previously classified as subspecies of B. fragilis; (2) the melaninogenicus-oralis group, which includes the three saccharolytic subspecies (ss.) of B. melaninogenicus--ss. melaninogenicus, ss. intermedius and ss. levii--and four non-pigmented species; (3) the asaccharolytic group, which comprises B. asaccharolyticus (formerly B. melaninogenicus ss. asaccharolyticus), B. corrodens and other non-pigmented non-saccharolytic strains, and (4) the fusobacteria.

The role of Robertson's cooked-meat broth in the bacteriological evaluation of surgical specimens.

The results were compared of submitting simple swabs, swabs in Stuart's Transport Medium (STM) and swabs in Robertson's cooked-meat broth (RCMB), from 100 potentially or definitely infected sites in patients undergoing general surgery. Significantly more positive bacterial cultures were obtained from swabs sent in RCMB (65), than from swabs sent either in STM (39) or as simple swabs (32). The isolation of potentially significant organisms from only the RCMB series could influence clinical management. The conventional reluctance of bacteriologists to accept evidence obtained from RCMB cultures seeded directly in the ward or at operation is challenged.