Detection of antigen in sera of patients with invasive aspergillosis: intra- and interlaboratory reproducibility. The Dutch Interuniversity Working Party for Invasive Mycoses.

The intra- and interlaboratory reproducibilities of a commercial sandwich enzyme-linked immunosorbent assay (ELISA) for the detection of Aspergillus galactomannan in serum (Platelia Aspergillus; Sanofi Diagnostics Pasteur, Marnes-La-Coquette, France) were evaluated in six laboratories of university hospitals. Twenty serum samples were obtained from 12 neutropenic patients including 6 with invasive aspergillosis. These samples were blinded and sent to each center together with eight blinded ELISA-negative serum samples spiked with known concentrations of galactomannan. The centers were provided with ELISA microtiter plates from a single batch and a detailed protocol. Ten clinical samples showed ELISA reactivity, while 10 samples were ELISA negative. The mean coefficient of variation (CV) of the optical density values was 4.24% within a single assay and 25.6% between runs. The interassay CV of the ratios for the serum samples tested was 18.6%. Analysis of ordinal interpretation of the ELISA result (i.e., negative, gray zone, or positive) showed excellent reproducibility. Recalculation of the cutoff values for positive and negative samples suggested that the cutoff level recommended by the manufacturer could be lowered from 1.0 to 0.8 for negative samples and from 1.5 to 1.0 for positive samples. The intra- and interlaboratory reproducibilities were excellent when the ELISA results were interpreted as ordinal data, but considerable variation in optical density values and, to a lesser extent, in the ratios for the serum samples tested, was observed between runs. High assay variability was also found for serum samples spiked with known concentrations of galactomannan. Therefore, antigen titers in serum samples from a single patient, measured in different runs, should be compared with caution.

Prevalence of penicillin-resistant viridans streptococci in healthy children and in patients with malignant haematological disorders.

The prevalence of penicillin-resistant viridans streptococci was studied in healthy children and in paediatric and adult patients with leukaemia to determine whether the frequent presence of penicillin-resistant streptococci in the oral cavity of children with leukaemia is the result of antibiotic therapy. Twenty of the oral swabs from 50 healthy children who had not received antibiotics in the three months prior to sampling yielded viridans streptococci that could be cultured on blood agar containing 2 micrograms/ml benzyl-penicillin. In 11 of the 20 cases the streptococci were resistant to penicillin (MIC > or = 4 micrograms/ml). This prevalence is significantly higher than that found in adult leukaemia patients (40% vs. < or = 5%) but is about the same as that found in paediatric patients with leukaemia. The high prevalence of penicillin-resistant streptococci in the paediatric age group should be considered when selecting therapy and prophylaxis, especially when the risk of infection with one of these cocci is enhanced.

Risk factors for fungal infection in patients with malignant hematologic disorders: implications for empirical therapy and prophylaxis.

To determine which patients are at high risk for disseminated fungal infection and should be given systemic prophylaxis, we studied the charts of 341 patients with malignant hematologic disorders who were admitted to our institution during 10 consecutive years. These patients represented 636 admissions; during these admissions, 60 invasive fungal infections occurred, with deaths in 44 cases. All patients who died of these infections either had persisting granulocytopenia and a poor prognosis for the underlying disease or suffered from chronic graft-vs.-host disease. Two of 58 patients who had no or low-level candidal colonization developed this infection (P < .001). Nine of the 10 patients with candidal infection had microbiologically proven bacteremia within the week preceding the candidal infection. After bone marrow transplantation, 8 of 10 patients with chronic graft-vs.-host disease vs. 2 of 36 without this disease (P < .001) developed fatal infection with Aspergillus species. The results of our study reveal that patients with high-level candidal colonization who were treated for microbiologically proven bacteremia and patients with chronic graft-vs.-host disease might benefit from systemic antifungal prophylaxis.

Selective decontamination in bone marrow transplant recipients.

Patients undergoing bone marrow transplantation become immunocompromised for various reasons. Deep granulocytopenia, induced by conditioning (chemotherapy and total body irradiation), renders the patient at risk for serious bacterial and fungal infections. Our strategy for prevention of these infections by selective decontamination (SD) is the result of more than 15 years of clinical experience and research. The combination of antibiotics, used as standard SD (neomycin, polymyxin B, pipemidic acid and amphotericin B), with the application of local antimicrobial agents eliminates aerobic Gram-negative rods, Staphylococcus aureus and Candida spp. from the mucosal surfaces of the digestive tract, while the majority of the anaerobic flora persist and support colonization resistance (CR). The antibiotics used either are not resorbed or do not yield therapeutic serum concentrations. Antibiotics which induce therapeutic serum concentrations, such as ciprofloxacin and cotrimoxazole, are only used for SD on a limited scale. When Gram-negative rods persist despite intake of the standard regimen, ciprofloxacin is given until these persisting rods are eliminated. If the patients cannot swallow the oral regimen, i.v. cotrimoxazole is given temporarily. Streptococcal infections are prevented by the i.v. administration of penicillin for 14 days starting on the first day after cytotoxic treatment (conditioning for bone marrow transplantation). The combination of SD and systemic prophylaxis has been shown to be adequate; the major problem then remaining is a relatively mild catheter-associated infection with coagulase-negative staphylococci.

Prevention of viridans-group streptococcal septicemia in oncohematologic patients: a controlled comparative study on the effect of penicillin G and cotrimoxazole.

In a controlled randomized study among 48 patients undergoing 75 courses of aggressive antileukemic therapy, it was shown that cotrimoxazole was less effective than penicillin G in preventing septicemia due to viridans streptococci. Both antibiotics were given intravenously. During 35 episodes of chemotherapy in the group of patients on penicillin G only, one patient developed a streptococcal bacteremia; this contrasted with bacteremia and septicemia in seven patients during 40 episodes in the group on cotrimoxazole. In three of these seven patients, septicemia was associated with respiratory failure and it was the cause of death in one. Both aerobic gram-negative rods and streptococci which caused infection despite cotrimoxazole prophylaxis were resistant to cotrimoxazole. Side effects such as hypersensitivity and favorable or unfavorable interaction with the oral selective decontamination regimen were similar for the two drugs, with the exception of colonization with Candida spp, which occurred more often in patients on cotrimoxazole than in patients on penicillin.

The relevance of cultures of catheter-drawn blood and heparin-lock fluid to diagnose infection in hematologic patients.

The results of bacteriologic cultures of blood and heparin-lock fluid, both drawn from the central venous catheters of 54 consecutive oncohematologic patients, have been used to determine their value for the diagnosis of systemic and catheter-associated infection. In 30 patients with clinical signs of infection (bacteremia or septicemia), 74 of 1000 (7.4%) heparin-lock fluid cultures, 114 of 542 (21%) catheter-drawn blood cultures, and 36 of 134 (26%) venipuncture blood cultures became positive, whereas in 24 patients without clinical signs of infection the respective values were 5 of 700 (0.7%), one of 220 (0.4%), and none of ten cultures. Comparison of the results of cultures sampled on the same day reveals that the positive and negative predictive values for catheter-drawn blood cultures, with the venipuncture blood cultures taken as the standard for bacteremia, are 82% and 95% respectively. The results of heparin-lock fluid are indicative for clinically relevant colonization of the catheter. Three or more positive heparin-lock fluid cultures, sampled on subsequent days, were correlated with the occurrence of bacteremia or septicemia with a positive predictive value of 100%. The conclusions are supported by the results of scanning electron microscopy.

Modulation of the intestinal flora of mice by parenteral treatment with broad-spectrum cephalosporins.

A study was performed to determine the effect of parenteral treatment with four broad-spectrum cephalosporins (cefoperazone, ceftriaxone, ceftazidime, and cefepime) on the number of aerobic gram-negative rods and on the outgrowth of Candida albicans and a multiresistant strain of Citrobacter freundii in the feces of mice. The estimated fractions of a parenteral dose that were excreted into the gastrointestinal tract were 0.37 for cefoperazone, 0.11 for ceftriaxone, 0.03 for ceftazidime, and 0.002 for cefepime. All four cephalosporins significantly decreased the number of aerobic gram-negative rods in the feces, and virtually all gram-negative rods were eliminated at high doses of cefoperazone, ceftazidime, and ceftriaxone. Furthermore, at high doses these three compounds led to a significant increase of the outgrowth of resistant Citrobacter freundii. The outgrowth of Candida albicans was increased at high doses of cefoperazone and ceftriaxone, whereas ceftazidime and cefepime did not have this effect. The most profound changes in the gastrointestinal ecology were observed during treatment with high doses of cefoperazone. The results suggest that the colonization resistance of the gastrointestinal tract can be substantially decreased by parenteral treatment with cefoperazone and, to a lesser extent, with ceftriaxone and ceftazidime.

Modulation of the intestinal flora of mice by treatment with aztreonam and tigemonam.

Oral and parenteral administration of aztreonam and oral administration of tigemonam to conventional mice caused a decrease in the number of aerobic gram-negative rods in the feces. Oral treatment with high doses of aztreonam (greater than or equal to 25 mg/kg/day) and tigemonam (100 mg/kg/day) adversely influenced colonization resistance, whereas oral treatment with lower doses of the monobactams or parenteral treatment with aztreonam did not.

Comparison of the effects of aztreonam and tigemonam against Escherichia coli and Klebsiella pneumoniae in vitro and in vivo.

A study was performed to investigate the pharmacodynamics of aztreonam and tigemonam against Escherichia coli and Klebsiella pneumoniae in vitro and in vivo. The in vitro concentration-effect relationships were determined in short-term growth experiments. The in vivo dose-effect relationships were determined in an experimental thigh muscle infection in irradiated mice. In this model, E. coli was injected into one thigh muscle and K. pneumoniae was injected into the other. Throughout these experiments aztreonam was administered subcutaneously and tigemonam was administered orally. For analysis of the antibacterial pharmacodynamics, the following parameters were determined: the maximum effect as a parameter for efficacy, the 50% effective concentration (or dose) as a parameter for potency, and the slope of the concentration-effect relationship. To assess the relationship between the concentration of the antibiotic and the antibacterial effect in vivo, the pharmacokinetics of the two drugs in the plasma of mice were determined as well. The maximum in vitro and in vivo effects of aztreonam and tigemonam against both bacteria did not differ substantially. However, both drugs killed E. coli more effectively than K. pneumoniae, indicating that the maximum in vitro effect of these drugs against E. coli was higher than that against K. pneumoniae. The maximum in vivo effect of both drugs against E. coli was similar to that against K. pneumoniae. Furthermore, in vitro aztreonam was about twice as potent as tigemonam, but in vivo the reverse was the case. These findings were explained by pharmacokinetic differences between subcutaneously administered aztreonam and orally administered tigemonam, because concentrations of tigemonam in plasma remained at microbiologically active concentrations longer than those of aztreonam did.

Isolation of vancomycin-resistant enterococci in haematologic patients.

After the occurrence of septicaemia with a vancomycin-resistant Enterococcus faecalis strain in a patient, it was decided to determine the number of carriers of vancomycin-resistant cocci among haematologic patients. During a period of six months 135 stool samples from 25 children, and 400 samples from 70 adults were studied. All samples from the children were negative for vancomycin-resistant cocci. Nine of the adult patients had cultures positive for cocci, all identified as enterococci, which were highly resistant to vancomycin (MIC greater than 250 micrograms/ml), sensitive to amoxicillin, moderately resistant to gentamicin, slightly resistant to teicoplanin, and sensitive to daptomycin. None of these patients had been given vancomycin prior to the isolation of the vancomycin-resistant enterococci.

Vancomycin-resistant Enterococcus faecalis in a bone-marrow transplant recipient.

Since 1988 a number of reports on the emergence of vancomycin-resistant enterococci have been published. We describe an additional case of colonization of and subsequent infection with a vancomycin-resistant Enterococcus faecalis in a bone-marrow transplant recipient, who had never before received vancomycin therapy. The strain was resistant to most antibiotics tested, including low-level resistance to gentamicin and cross-resistance to teicoplanin. It was sensitive to amoxicillin and the lipopeptide antibiotic daptomycin. The origin, the mode of acquisition and the incidence of vancomycin-resistant enterococci are still unknown. The clinical and bacteriological features of the cases reported thus far and the genetic and biochemical basis of vancomycin resistance are discussed.

Acquired severe aplastic anaemia in adults--a single centre study with 13 years follow-up.

Between 1974 and July 1987 the diagnosis of severe aplastic anaemia (SAA) was confirmed in 82 patients. Overall actuarial survival was 57% at 7 yr. Four patients recovered while receiving conventional therapy, and four died before treatment with antithymocyte globulin (ATG) or bone marrow transplantation (BMT) could be initiated. Nineteen patients (median age 19.6 yr) were treated with allogeneic BMT (11 as initial therapy, eight after ATG). Incidence of acute and chronic graft versus host disease was high, occurring in 14/16 and 4/11 patients at risk, respectively. Survival of BMT patients (18/19 transfused) was 32% at 7 yr. Of 63 patients treated with ATG, survival was 63% at 7 yr but decreased to 43% at 11 yr. The 2.5 yr survival following ATG was influenced by pretreatment disease severity (defined by percentage reticulocytes, granulocyte and platelet counts), age and--in patients under 45 yr of age--by sex. However, pretreatment disease severity was less in patients aged between 20 and 45 yr and in females. Concomitant androgen therapy, animal source of ATG, interval diagnosis--ATG (which was in general rather short) and aetiology did not influence survival. Thirty-four patients became transfusion independent for up to 26 months after ATG. A gradual increase in granulocyte and platelet counts could be observed over a period of many years, and 26 patients recovered to show a normal haemoglobin level, granulocytes greater than or equal to 1.0 X 10(9)/l and platelets greater than or equal to 100 X 10(9)/l). Late complications (paroxysmal nocturnal haemoglobinuria, myelodysplastic syndrome/acute leukaemia, hepatocellular carcinoma) were observed in nine patients who survived with autologous marrow function. Five died within 12 yr of initial therapy.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparative study of the effects of four cephalosporins against Escherichia coli in vitro and in vivo.

A thigh muscle infection induced with Escherichia coli in irradiated mice was used as a model to compare the in vivo pharmacodynamics of the antibacterial effect of four cephalosporins (i.e., cefepime, ceftriaxone, ceftazidime, and cefoperazone) with the in vitro antibacterial pharmacodynamics of these drugs. The following in vitro pharmacodynamic parameters were determined: the maximum effect as a measure for efficacy, the 50% effective concentration as a parameter for potency, and the slope of the concentration-effect relationship. For analysis of the in vivo antibacterial pharmacodynamics, the same parameters were applied for the dose instead of the concentration. For the detection of a relationship between concentration and antibacterial effect in vivo, we determined the pharmacokinetics of the four cephalosporins in the plasma of mice. The results showed that, in general, there is a direct relationship between the in vivo and in vitro pharmacodynamics of these cephalosporins. The maximum effects of cefepime, ceftazidime, and cefoperazone were approximately similar in vivo and in vitro. The sequence of potency of these drugs was, in descending order, cefepime, ceftazidime, and cefoperazone. Ceftriaxone differed from the other three cephalosporins in that it displayed unexpected in vivo pharmacodynamics. Ceftriaxone was just as efficacious as the other three in vitro, but its maximum effect in vivo was much lower. This relatively low maximum effect of ceftriaxone in vivo was not explained by the pharmacokinetic characteristics of the drug. From the present results it can be concluded that the in vitro efficacy of cephalosporins does not necessarily have a predictive value for the in vivo efficacy.

Relation between the therapeutic efficacy of imipenem in an experimental infection in granulocytopenic mice and its effect on murine intestinal microbial ecology.

A study was performed to assess the effect of parenteral administration of imipenem on the intestinal microbial ecology of mice in relation to the therapeutic efficacy of the drug. The therapeutic effect of imipenem was assessed in a short-term experimental thigh muscle infection with Escherichia coli in irradiated mice. The maximum antibacterial effect of imipenem was approached at a single dose of 80 mg/kg. At this dose of imipenem the number of viable E. coli in the thigh muscle decreased from 9 x 10(6) to 4 x 10(5), whereas in untreated controls this number increased from 9 x 10(6) to 2 x 10(8). Daily administration of a dose of 80 mg/kg imipenem caused significant changes in the intestinal flora of conventional mice, despite the low concentrations of the drug (about 1 mg/kg) found in the caecum 30 min after infection of this dose. These changes consisted of an increase in the number of E. coli and enterococci and a decrease in the number of Staphylococcus aureus in the faeces. Furthermore, at this dose of imipenem, the number of Candida albicans recovered from the faeces after oral contamination with this organism was increased. We conclude from these animal experiments that the use of imipenem in high doses causes ecological changes in the intestinal bacterial flora with the potential to promote colonization by candida.

Respiratory failure elicited by streptococcal septicaemia in patients treated with cytosine arabinoside, and its prevention by penicillin.

In order to prevent septicaemia with streptococci, 20 consecutive selectively decontaminated patients on intermediate high-dose Ara-C treatment for malignant haematological diseases were given penicillin G. The incidence of infection with streptococci decreased from 0.76 per episode (14 patients, 17 episodes) for controls who did not receive penicillin G to 0.11 per episode in the prophylaxis group (20 patients, 26 episodes). Simultaneously, a decrease in the incidence of respiratory failure was observed, i.e. 0.52 per episode for controls and 0.19 per episode for patients on penicillin G. The results suggest that septicaemia with streptococci triggers the development of noncardiogenic pulmonary oedema in patients with pre-existing damage of the lung due to aggressive cytotoxic treatment. This suggestion is supported by the sequence of events, regarding the occurrence of infection and respiratory failure and the results of measurements of antileukoprotease serum concentrations, a parameter for pulmonary capillary leakage. Taking into account the data in the literature and the results of the present study, the conclusion is drawn that in patients treated with (intermediate) high dose Ara-C, prevention of streptococcal septicaemia is associated with a decrease in the incidence of respiratory failure.

Influence of cytostatic treatment on the efficacy of erythromycin and roxithromycin in a staphylococcal infection in mice.

Mice made monocytopenic with etoposide or both granulocytopenic and monocytopenic with cyclophosphamide were infected in a thigh muscle with 3 x 10(6) CFU of Staphylococcus aureus; 1 h later erythromycin or roxithromycin was administered, and 4 h after that the number of CFU per thigh was determined. In vitro as well as in vivo, the maximal effect of both antibiotics was only bacteriostatic. Monocytopenia did not diminish the efficacy of either erythromycin or roxithromycin in vivo, whereas the combination of granulocytopenia and monocytopenia markedly decreased the efficacy of both drugs: a 4-fold dose increase was necessary to obtain the same final number of CFU at the site of infection as in the controls. It is concluded that granulocytes contributed substantially to antibiotic efficacy against S. aureus in this short-term infection model.

Decontaminating efficacy of ciprofloxacin in an animal model.

A study was performed to assess the suppressive effect of ciprofloxacin on the aerobic gram-negative flora of the gastrointestinal tract in mice in relation to its effect on colonization resistance. Ciprofloxacin exhibited a potent decontaminating effect, with the maximum effect being reached at a dose of 8 mg/kg/day. At this dose aerobic gram-negative rods were eliminated from the gastrointestinal tract within 24h without any adverse influence on the parameters of colonization resistance measured.