W. A. C. Vel

KILLING CAPACITY OF HUMAN POLYMORPHONUCLEAR LEUKOCYTES IN AEROBIC AND ANAEROBIC CONDITIONS

The killing by human polymorphonuclear leukocytes of several species of bacteria, some of which were catalase positive, was examined in vitro in aerobic and anaerobic conditions. When all conditions other than the oxygen tension were identical, killing after 30 min was slightly greater in aerobic than in anaerobic conditions. However, after 60 and 120 min the difference between aerobic and anaerobic killing was smaller, and killing was nearly complete for all strains tested. These results conflict with the common opinion that oxygen is essential for efficient killing. Minor differences in experimental conditions can greatly influence results, and may be responsible for the discrepancy between this study and some previous studies on this subject.

POLYMORPHONUCLEAR LEUKOCYTE CHEMOTAXIS BY MIXED ANAEROBIC AND AEROBIC BACTERIA

The induction of chemotactic activity of polymorphonuclear leukocytes (PMNL) by anaerobic and aerobic bacteria alone or in combination was evaluated. Washed cells as well as the supernate of Proteus mirabilis were chemotactic for leukocytes. The supernate of cultures of two strains of Bacteroides fragilis contained small amounts of chemotactic factors. No chemotactic factors were released from the non-fragilis Bacteroides strains. The supernates of cultures of anaerobic bacteria were capable of inhibiting chemotaxis of leukocytes to the chemotactic factors of P. mirabilis. P. mirabilis and two strains of B. fragilis generated chemotactic factors in serum but none of the other Bacteroides spp. tested were able to induce serum chemotactic factors.

Pathogenic synergy between Escherichia coli and Bacteroides fragilis or B. vulgatus in experimental infections: a non-specific phenomenon.

The virulence of Bacteroides fragilis and B. vulgatus for mice was compared in a skin-infection model. These strains were also tested for pathogenic synergy in mixed infections with Escherichia coli. Strains of B. fragilis were generally more virulent than strains of B. vulgatus and, with one exception, the effect of Bacteroides strains in mixed infections merely reflected their inherent virulence.

Pathogenic synergy between Escherichia coli and Bacteroides fragilis: studies in an experimental mouse model

An animal model is described for quantitative evaluation of pathogenic synergy between Escherichia coli and Bacteroides fragilis in which adjuvants were not required for abscess formation. Two sets of strains of E. coli and B. fragilis isolated from human wound infections were tested. Pathogenic synergy was observed in only one of the two combinations and was dependent on properties of E. coli.

Pathogenic synergy: mixed intra-abdominal infections

In this article we review our researches into the pathogenesis of mixed infections. These may conveniently be divided into in vitro and in vivo studies.In vitro we confirmed that interference with the killing of aerobes by polymorphonuclear leucocytes (PMN’s) is a property of theBacteroides strains tested and appears to depend on competition for opsonins i.e. complement factors. Further studies are in progress to define which complement factors and which bacterial structures are involved. The influence ofB. fragilis on chemotaxis has also been studied. Our preliminary data suggest thatB. fragilis is itself poorly chemotactic and reduces the chemoattractivity ofProteus mirabilis. This observation is surprising when we consider that abscess formation is the hall-mark ofB. fragilis infections and needs clarification.In vivo we have developed a skin infection model in mice which is economical and gives reproducible and quantitative results. In this model we have demonstrated pathogenic synergy betweenEscherichia coli andB. fragilis. Further studies are planned to assess the role of complement and bacterial factors in this in vivo synergy.

Killing of Escherichia coli by human polymorphonuclear leucocytes in the presence of Bacteroides fragilis.

The inhibitory effect of Bacteroides fragilis on the in vitro killing of Escherichia coli by polymorphonuclear leucocytes was studied with two pairs of E coli and B fragilis isolated from human wound infections. Both B fragilis strains behaved similarly: they inhibited the killing of one E coli strain, while the killing of the other E coli strain was not affected. The different behaviour of the two E coli strains depended on their need for fresh serum in the killing by the polymorphonuclear leucocytes. The inhibitory effect of the B fragilis strains could be completely accounted for by their effect on complement.

Effect ofBacteroides fragilis grown in the presence of clindamycin, metronidazole and fusidic acid on opsonization and killing ofEscherichia coli

Bactericidal action of human polymorphonuclear leucocytes onEscherichia coliin the presence ofBacteroides fragilisgrown in subinhibitory concentrations of clindamycin, metronidazole and fusidic acid was studied.Bacteroides fragilisgrown in the absence of drugs significantly inhibited the killing ofEscherichia coli.Bacteroides fragilis grown in the presence of the drugs had a reduced inhibitory effect on the killing ofEscherichia colibut this reduction was only significant forBacteroides fragilisgrown in 1/2 MIC of clindamycin. The phagocytosis ofBacteroides fragilisgrown with and without clindamycin, as measured by killing, was the same. Complement consumption ofBacteroides fragilisgrown with and without clindamycin did not differ. Clindamycin-treatedBacteroides fragilisfixed C3 to a significantly lower degree than did untreated bacteria. The chemiluminescence ofEscherichia coliopsonized with serum preincubated with clindamycin-treatedBacteroides fragiliswas significantly higher than with serum preincubated with untreated bacteria. These results suggested that in killing experiments of mixedEscherichia coliandBacteroides fragilis,the mechanism underlying the reduced inhibitory capacity of clindamycin-exposedBacteroides fragilisis related to greater availability of C3 in serum for opsonization ofEscherichia coli.

Interactions between polymorphonuclear leucocytes, Bacteroides sp, and Escherichia coli: their role in the pathogenesis of mixed infection.

Five Bacteroides fragilis strains and five Bacteroides vulgatus strains were compared with regard to their ability to consume complement and to fix C3, their killing by polymorphonuclear leucocytes, and their ability to inhibit the bactericidal effect of serum and polymorphs on Escherichia coli strains. Complement consumption was positively related to C3 fixation, but no relation was observed between these variables and the killing of the anaerobes. Greatest inhibition of the killing of E coli by serum and polymorphs was achieved with the bacteroides strains that fixed most complement. The greater virulence of B fragilis in mixed infections with E coli was not reflected either by a greater ability to inhibit the killing of E coli or a greater resistance of the anaerobes themselves to the bactericidal effect of serum and polymorphs.

The effect of antimicrobial agents on the killing of Escherichia coli in the presence of Bacteroides fragilis

Bactericidal activity of human polymorphonuclear (PMN) leucocytes on Escherichia coli in the presence of Bacteroidesfragilis grown in sub-inhibitory concentrations of clindamycin, metronidazole and fusidic acid was studied. B.fragilis, grown in the absence of antibiotics, significantly inhibited the killing ofE. coli. When B.fragilis was grown in the presence of sub-inhibitory concentrations of antibiotics its inhibitory effect on the killing of E. coli was reduced. In spite of a positive effect of all three drugs only B.fragilis grown in sub-inhibitory concentrations ofclindamycin had a significantly reduced inhibitory effect on PMN leucocyte killing of E. coli. Killing, chemiluminescence and complement consumption of B.fragilis grown with and without clindamycin were not different. Clindamycin-treated B.fragilis fixed C3 to a significantly lower degree than did untreated bacteria. These results may have clinical relevance because during the antibiotic therapy there will be often periods when only sub-inhibitory concentrations of the antimicrobial are present.

Bactericidal activity of polymorphonuclear leukocytes in aerobic and anaerobic conditions

Since the introduction of hepatitis B vaccines indications for vaccination are discussed. The New York study on homosexual men (Szmuness et al., 1980) clearly showed a high grade of protection against infection with hepatitis B virus. The same appeared to be the case in trials in dialysis units. An advisory committee of the Netherlands Health Council raised the question which groups in the community have to be considered for vaccination. Two categories were identified. The first consists of patients who, because of special circumstances, are more susceptible than average to infection with hepatitis B virus. The second category consists of healthy persons who, because of special circumstances, are constantly or occasionally exposed to possible infection, either in a medical setting or otherwise. To determine the order of priority for the second category criteria have to be set. An important tool for this procedure is the screening of the different subgroups on antibodies to HBsAg. This policy combines the possibilities of gaining insight into the attack rate of the virus in these subgroups and of setting up the priorities concerning vaccination. Screening of HBsAg has to be restricted to identify persons who when positive will endanger other individuals.