Computer processing of microscopic images of bacteria: morphometry and fluorimetry.

Several techniques that use computer analysis of microscopic images have been developed to study the complicated microbial flora in the human intestine, including measuring the shape and fluorescence intensity of bacteria. These techniques allow rapid assessment of changes in the intestinal flora and could apply equally to other complex microbial ecosystems.

A novel technique for detecting IgA coated potentially pathogenic microorganisms in the human intestine.

This paper describes a novel method for detecting immunoglobulin A (IgA) coated potentially pathogenic microorganisms (PPMs) in the human intestine. Essentially, the technique consists of 2 phases: one in which IgA coated bacteria are detected by immunofluorescence and a second in which these bacteria are subcultured in situ and subsequently identified. In this way transient bacteria are differentiated from resident bacteria. These results show that the resident bacteria are coated with IgA. Resident microorganisms are always highly concentrated in the digestive tract. These results strengthen the hypothesis that only the high antigen concentrations achieved by a large number of resident bacteria are capable of IgA induction.

Influence of temocillin on colonisation resistance and consequences for therapy.

The influence of temocillin on the colonisation resistance of the digestive tract was studied in mice, with doses ranging from 0.062 to 1 mg/day administered orally and 0.25 to 4 mg/day administered subcutaneously. The effect on the colonisation resistance was measured by the concentration of resistant bacteria (either endogenous enterococci or an exogenous strain of Enterobacter cloacae) as well as by the relative caecal weight and the appearance of beta-aspartylglycine in the caecal contents. None of these parameters indicated a decreased colonisation resistance by any of the dosages investigated, meaning that, in the mouse the drug can be used safely without an increased risk of overgrowth by resistant bacteria causing superinfections. Both systemic and oral administration of 0.25mg of temocillin/mouse/day (10 mg/kg/day) appeared to completely suppress the sensitive Gram-negative bacilli from the digestive tract within 2 days. This selective decontamination of the digestive tract in mice was the reason for a subsequent study in man. It emerged that selective decontamination with parenterally administered temocillin was possible in 7 out of 10 volunteers. The remaining 3 however, did not respond to temocillin although their faecal Gram-negative bacilli were susceptible to the drug. In none of the volunteers did the colonisation resistance appear to be affected. The consequences of the use of temocillin for either prevention or therapy of infections are discussed.

Influence of high versus low intestinal concentration of gram-negative bacteria and endotoxin on the susceptibility of murine myelopoiesis in bone marrow and spleen to cytostatic treatment with Ara-C.

The haematopoietic recovery after i.v. cytarabine was studied in C3H/Law mice as a measure for stem cell susceptibility in relation to the intestinal Gram-negative bacteria (GNB) and endotoxin. Reduction or elevation of GNB and endotoxin was induced by either polymyxin or bacitracin, both non-absorbable antibiotics. Bacitracin caused less suppression of the splenic cellularity after cytarabine, and an advancement of the recovery of femoral nucleated cells. The femoral recovery of CFU-GM exhibited a biphasic pattern. The speed and height of the rebound increase of CFU-GMs were significantly affected by the antibiotics. Thus, (modulation of) the murine intestinal microflora influences the haematopoietic recovery after cytostatic drugs. The mechanisms involved are complex; intestinal endotoxin seems to play a role.

Recovery of murine myelopoiesis after cytostatic reduction by Ara-C. Effect of bacitracin-induced changes in the intestinal microflora and influence of timing.

The influence of intestinal flora modulation by oral bacitracin on the recovery of myelopoiesis after Ara-C was studied in C3H/Law mice. Bacitracin resulted in a 3-5 log increase of Gram-negative bacteria and a 10-fold increase of the intestinal endotoxin concentration. Initiation of bacitracin before Ara-C stimulated the initial rebound increase of colony-forming units for granulocytes and macrophages (CFU-GM) from 23.2 +/- 1.3 to 28.4 +/- 1.4 x 10(3) per femur. Starting the bacitracin after Ara-C advanced the second phase of the rebound CFU-GM increase with 6 days. An important role in the recovery of myelopoiesis after cytostatic drugs in C3H/Law mice is suggested for the intestinal Gram-negative microflora, probably mediated by bacterial endotoxin.

Modulation of the intestinal microflora by the non-absorbable antibiotic vancomycin leads to a reduction of the tumor load in liver and spleen in a leukemic rat model.

Based on previous studies where it was shown that non-absorbable antibiotics can influence the normal hematopoiesis via changes in factors related to the intestinal microflora, the influence of vancomycin on the progression of acute myeloid leukemia was investigated in the BNML rat model. Oral vancomycin, which selectively reduces Gram-positive bacteria in the gut, leads to diminution of the leukemic load in liver and spleen by 30-60%. This 'antileukemic effect' is not dependent on Gram-negative bacteria as source for endotoxin. The presumed mechanism is a decrease of the leukemic growth fraction caused by alterations in the absorption of substances from intestinal Gram-positive bacteria.

Acinetobacter mediastinitis and pneumonia in a thorotrastoma patient. The oropharyngeal flora as source of infection.

A case of bilateral pneumonia, mediastinitis and septicaemia caused by Acinetobacter calcoaceticus and Candida albicans is described. The infections occurred after a palliative operation for an esophagotracheal fistula in a thorotrastoma patient. The oropharynx was colonized by the two microorganisms at admission and is presented as the source of these infections. Clinical management and antimicrobial policy, including oropharyngeal decontamination, leading to a good outcome are reported.

Clinical use of selective decontamination: the concept.

Infections can be classified according to: (1) the type of offending microorganism (virus, bacteria, fungi, parasites), (2) according to the clearance by the defence system (T cell dependent/independent) and (3) in case bacteria are the causative agents in Gram-positive and Gram-negative infections. The latter classification in Gram-positive and Gram-negative infections has appeared to have a practical consequence. Gram-negative bacteria, often involved in major infections and yeasts, appear to play practically no role in the intestinal ecological system. Consequently, it is nowadays increasingly attempted to eliminate Gram-negative bacteria and yeasts selectively from the digestive tract with antimicrobial agents. Selective suppression of Gram-positive bacteria may severely affect the ecosystem of the digestive tract. This selective suppression of Gram-negatives must be continued as long as patients are immunocompromised (locally or systemically) and is called selective decontamination of the digestive tract.

Development and persistence of multi-resistance to antibiotics in bacteria; an analysis and a new approach to this urgent problem.

The intestinal microflora may have more influence on infectious diseases, than the mere control of growth of opportunistic micro-organisms by colonisation resistance (CR) and unspecific stimulation of the immune system. In compromised patients the CR may become decreased for several reasons but mostly because antibiotics reach the intestine during treatment. The consequence of a CR-decrease is that antibiotic-resistant opportunistic micro-organisms may increase in numbers in the gut. In this context, it is hypothesised that if the CR could be maintained at a normal level, the risk for maintenance and spread of resistant strains could be mitigated. Such maintenance requires absence of active antibiotic substance in the gut. This might be brought by the inactivation of antimicrobial agents by intestinal contents. Intra-intestinal inactivation has been described to occur along two possible routes: (1) inactivation by chemical binding or absorption and (2) by enzymatic destruction. Secondly, the composition of the intestinal microflora should be maintained at a normal level in case of other reasons for CR-decrease than antibiotic activity. Comprehensive study of the composition of normal microflora and the strains of species which play a role in CR with techniques which have become available during last decade, is recommended as well as the application of certain pre- and probiotics. It is concluded that antibiotic inactivation may be an ancient strategy of nature which should become incorporated in antibiotic treatment. Antibiotic use and development of resistance may have occurred when ecosystems formed several billions of years ago. Protection against antibiotics produced by newcomers into the ecosystem may have developed as it was necessary to maintain locally available nutrients for the inhabitants of the ecosystem. Should this hypothesis be correct, it is plausible that antimicrobial inactivation by antibiotic inactivating molecules is ubiquitous. In the ecosystem of the digestive tract, molecules involved in inactivation may predominantly be formed by microorganisms.

Selective decontamination of the digestive tract of Syrian hamsters.

Conventional Syrian hamsters colonized with aerobic gram-negative bacteria such as Pasteurella pneumotropica and various Enterobacteriaceae species were successfully and permanently freed from these microorganisms by oral treatment for 4 weeks with dihydrostreptomycin and 'Orabase' premixed with appropriate antibiotics. Concomitant oral treatment with dimetridazol for the elimination of intestinal flagellates was unsuccessful. During treatment the animals were maintained under germ-free isolation conditions.