Diagnostic testing for Legionnaires' disease in the Netherlands between 2007 and 2009: a possible cause for the decline in reported Legionnaires' disease patients.

Legionnaires' disease (LD) is an acute pneumonia caused by the inhalation or aspiration of aerosols contaminated with the Legionella bacteria. In the Netherlands, around 300 LD cases per year were reported between 2000 and 2008, but in 2009, the number dropped to 251, which was the lowest number in the previous 5 years of surveillance. We investigated if this decrease could be explained by the number of performed Legionella diagnostic tests in this year. We analyzed the number of tests performed between 2007 and 2009 in three large microbiological laboratories in different geographical regions in the Netherlands. Our data showed that there was no decrease in the number of patients for whom a diagnostic test for Legionella was performed in this period. These results are not in line with our hypothesis that the decrease in reported Legionella pneumonia patients in 2009 would be due to a decrease in patients for whom a diagnostic test was performed. We conclude that it is more likely that other factors such as the influence of weather patterns might explain the sudden drop in reported Legionella pneumonia patients in 2009 compared to the previous years, and it would be interesting to investigate this for the period described.

[Recurrence of Clostridium difficile-associated diarrhoea prevented by the administration of a whey concentrate from specifically immunised cows; prospective study]. / Recidieven van Clostridium difficile-geassocieerde diarree voorkómen door toediening van een weiconcentraat van specifiek geïmmuniseerde koeien; prospectief onderzoek.

OBJECTIVE: To try to prevent recurrences of Clostridium difficile-associated diarrhoea (CDAD) by treatment with a specific neutralising secretory IgA-enriched whey-protein concentrate (40%) made from the milk of cows immunised with C. difficile and its toxins. DESIGN: Prospective, non-blinded, clinical cohort study. METHOD: In 2005-2006, 100 consecutive patients with CDAD received the whey concentrate for 2 weeks after completion of standard antibiotic therapy. For a period of 60 days after the start of the administration, the safety and preliminary efficacy of the whey concentrate were evaluated by means of a diary, blood determinations, active surveillance for adverse events, and the recurrence of CDAD. RESULTS: The whey concentrate was well tolerated and no safety issues were raised. Eleven out of 109 episodes (10%) were followed by a recurrence. After completion of the whey concentrate therapy, a positive test for faecal toxins or culture of C. difficile was predictive for the recurrence of CDAD (relative risk: 8.2 (95% CI: 1.04-64), and 4.7 (95% CI: 0.5-47), respectively). A positive faeces toxin during administration of the whey concentrate was also associated with an early recurrence of CDAD. CONCLUSION: Compared to historical and contemporary findings in control groups, the whey concentrate appeared to reduce the recurrence of CDAD by about 50%. However, the standard dose of the whey concentrate was probably not sufficient to fully neutralise the C. difficile toxins in faeces in all episodes.

Mycobacterium avium intracellulare otomastoiditis: case report and literature review.

Otomastoiditis caused by Mycobacterium avium intracellulare (MAI) is rare. Sub-optimal management of this condition can lead to significant morbidity and serious damage to the middle ear. Diagnosis is difficult, especially since most physicians are not familiar with the mode of presentation and symptoms. Presented here is a new case, followed by a review of the literature on MAI mastoiditis.

Bordetella pertussis and mixed infections.

AIM: In pertussis-like respiratory infections, once pertussis has been laboratory confirmed, other potential causative pathogens will seldom be looked for. Probably most mixed infections are found accidentally and since these mixed infections might cause a more severe disease we performed a retrospective study of their incidence. METHODS: We selected from 2 groups of patients with serologically confirmed Bordetella (B.) pertussis infection those in whom serology for other respiratory pathogens had also been performed. Group 1 consisted of 50 pertussis patients with 51 episodes of B. pertussis infection selected from 100 patients with serologically confirmed pertussis. They participated in a long-term follow-up after a B. pertussis infection. In group 2, 31 pertussis patients were selected from 98 consecutive patients with positive pertussis serology from one routine practice. RESULTS: In 23 of 82 pertussis infections (28%) serological evidence of 1 (n = 21) or 2 (n = 2) additional infections were demonstrated. These involved para-influenza virus (n = 6), respiratory syncytial virus (RSV) (n = 6), Mycoplasma pneumoniae (n = 5), adenovirus (n = 4), influenza A virus (n = 3) and influenza B virus (n = 1). CONCLUSIONS: We conclude that in patients with B. pertussis infection, coinfection with another respiratory pathogen is often present.

Community-acquired pathogens associated with prolonged coughing in children: a prospective cohort study.

A 2-year prospective study was performed of children with prolonged coughing to investigate the frequency of different respiratory pathogens, the rate of mixed infections, and possible differences in severity of disease between single and mixed infections. Sera from 135 children (136 episodes of prolonged coughing lasting 1-6 weeks) were tested for antibodies to different viruses and bacteria. Swabs were taken for culture and PCR to detect different viral and bacterial pathogens. One or more pathogens were found in 91 (67%) patients. One infectious agent was found in 49 (36%) patients, two agents in 35 (26%) patients, and more than two agents in seven (5%) patients. The most frequent pathogens encountered were rhinovirus (n = 43; 32%), Bordetella pertussis (n = 23; 17%) and respiratory syncytial virus (n = 15; 11%). The most frequent mixed infection was B. pertussis and rhinovirus (n = 14; 10%). No significant differences in clinical symptoms were observed between patients with or without pathogens; however, patients with mixed infections were significantly older. There was a strong seasonal influence on the number of infections, but not on the number of mixed infections. In children with prolonged coughing, there was a high frequency of mixed infections regardless of the season. However, mixed infection was not associated with increased disease severity. No clinical symptoms were found that allowed discrimination between specific pathogens.

In vivo uptake of daunorubicin by acute myeloid leukemia (AML) cells measured by flow cytometry.

Monitoring of daunorubicin (DNR) concentrations in leukemic cells in blood and bone marrow in vivo of patients with acute myeloid leukemia may yield insight into the interindividual variations of the clinical response to treatment. We evaluated the applicability of flow cytometry for measuring DNR uptake in direct comparison with high performance liquid chromatography (HPLC). In vitro studies revealed good correlations between the mean cellular fluorescence measured by flow cytometry and the cellular DNR concentrations determined with HPLC. In vivo cell measurements were then obtained in 17 evaluable patients during their first remission induction treatment with DNR and cytosine arabinoside. The results indicate that: (a) DNR fluorescence of leukemic blast cells is intermediate between the smaller lymphocytes and the approximately equally large granulocytes; (b) DNR fluorescence of peripheral blast cells and bone marrow blast cells correlate well (p less than 0.001); and (c) patients reaching complete remission show a tendency of higher DNR fluorescence of leukemic blast cells than do partial responders.

Cellular pharmacokinetics of daunorubicin: relationships with the response to treatment in patients with acute myeloid leukemia.

In an attempt to identify pharmacokinetic factors that determine the response of acute myeloid leukemia (AML) patients to induction chemotherapy, we determined the concentrations of daunorubicin (DNR) and the main metabolite daunorubicinol (DOL) in vivo and particularly evaluated the concentrations in blood and bone marrow nucleated cells. Cell measurements were obtained in 37 evaluable patients during their first remission induction treatment with DNR and cytarabine (ara-C) and directly compared with the plasma distribution kinetics of DNR. We show that (1) plasma DNR concentrations do not correlate with DNR concentrations in bone marrow nucleated cells; but (2) plasma area under the curve (AUC) values of DNR correlate inversely (P less than .01) with AUC values of DNR in WBCs; (3) concentrations of DNR in WBCs correlate positively (P less than .01) with DNR concentrations in bone marrow nucleated cells; and (4) the concentrations of DNR in WBCs show a negative correlation (P less than .01) with the numbers of peripheral blast cells at diagnosis. We then tested whether the pharmacokinetic parameters had predictive value for the clinical outcome of therapy, but none of the plasma levels or WBC and bone marrow concentrations of DNR predicted treatment outcome. The inverse correlation between the concentrations of DNR in WBC and the numbers of peripheral blast cells suggests that the effective DNR concentrations achieved intracellularly are mainly a function of the tumor load so that lesser amounts of DNR accumulate intracellularly when the AML cell numbers in blood are higher.

Concentrations of ciclosporin in allogeneic bone marrow recipients. Comparison of assay methods.

Thirteen patients in complete remission from acute nonlymphoblastic leukaemia or in chronic phase of chronic myelocytic leukaemia were treated with total body irradiation, cyclophosphamide and allogeneic bone marrow transplantation (BMT). Ciclosporin (CS) was administered for the prevention and the treatment of Graft versus Host Disease. Blood concentrations of CS were determined by Radioimmunoassay (RIA) and by High Performance Liquid Chromatography (HPLC). Trough levels of CS in peripheral blood as measured by RIA exceeded HPLC derived levels in nearly all (56/58) samples with a ratio of RIA:HPLC ranging from 2.43 +/- 1.42 at day 12 to 3.65 +/- 1.86 at day 26 after BMT (means +/- SD). A comparable ratio was found as regards the peak concentrations of CS in peripheral blood. Neither the dose of CS (0.5-3.0 mg/kg/day intravenously; 3.0-5.0 mg/kg/day per os) nor the duration of treatment (12, 19, 26 or 33 days after start of CS) were a significant factor as regards the ratio between HPLC and RIA. Concentrations of CS were also determined in bone marrow nucleated cells at 1 hour after the drug infusion had started. Here the ratio of RIA versus HPLC varied upon the duration of CS treatment with a highest ratio of 8.75 +/- 8.74 at day 12 after BMT. Bone marrow levels corresponded well with blood trough concentrations (p less than 0.01). It is concluded that the concentrations of CS in blood and bone marrow as determined by RIA and HPLC differ significantly, though consistently. At present, no advantage can be attributed to either method of analysis for routine clinical monitoring, as long as detailed information on the immunosuppressive and the toxic characteristics of CS metabolites in humans is lacking.

Kinetics of daunomycin in leukemia cells and leukocytes in vivo in the rat.

The capability of nucleated blood cells and leukemic cells to transport daunomycin (DNR) to target tissues in the body was investigated in the rat. The in vivo distribution kinetics of DNR entrapped in leukemia cells (brown Norway acute myeloid leukemia, BNML) or in nucleated bone marrow cells, which had been exposed to DNR (0.2 mg/5 X 10(7) cells) in vitro, were determined. It was found that BNML leukemia cells and normal bone marrow cells take up DNR according to a linear pattern up to 400 micrograms/5 X 10(7) cells. When these DNR loaded cells are infused into the rat, dose dependent distribution kinetics are observed. Compared to i.v. injection of the same dosage, cell-bound DNR leads to a higher concentration and a higher tissue area under the curve of DNR in the liver (P less than 0.05) and the spleen (P less than 0.05), while equal levels are attained in bone marrow. Lower concentrations and area under the curve of DNR are observed in cardiac tissue of normal rats (P less than 0.001) and leukemic rats (P less than 0.05). It is concluded that DNR entrapped into marrow and leukemia cells follows different kinetics from free DNR in plasma.

Pharmacokinetics of daunorubicin as a determinant of response in acute myeloid leukemia.

Twenty-one adult patients with acute myeloid leukemia (AML) were treated with the EORTC LAM-6 remission induction protocol [daunorubicin (DNR) (45 mg/m2, days 1-3), cytarabine (200 mg/m2, days 1-7) and vincristine (1 mg/m2, day 2)]. Pharmacokinetics of DNR were studied at day 1. The concentration of DNR and daunorubicinol were determined in plasma, in white blood cells and in bone marrow. A large variability was observed with respect to (1) the plasma area under the curve (AUC) 0-24 h (range: 0.06-0.37 nmol X h/ml); (2) the white cell AUC 0-24 h (range: 0-441 nmol X h/10(9) cells); and (3) the 1 h bone marrow concentration (range: 0-27 nmol/10(9) cells). In eight patients treated twice, a small intraindividual variability of these parameters was observed. Concentrations in plasma did not correlate with cellular concentrations. All pharmacokinetic parameters in plasma and white cells did not correlate with response to therapy. In patients reaching complete remission (CR), however, the tumor load, as expressed by the number of blast cells present in the untreated bone marrow, was significantly lower than the number of blast cells in patients not reaching CR.

Quantitative evaluation of intracellular uptake of daunorubicin in acute myeloid leukemia: a method analysis.

Critical technical parameters to establish a reliable method for quantifying the intracellular content of anthracyclines were evaluated in patients with acute myelocytic leukemia (AML); two methods were used for the isolation of leukocytes from the peripheral blood and two methods, for the extraction of daunorubicin (DNR), daunorubicinol (DOL), and doxorubicin (DOX) from these cells, followed by drug analysis using high-performance liquid chromatography (HPLC). At 0-4 degrees C the recovery of leukocytes after methylcellulose separation was low (64%). Cold hypotonic lysis gave better recovery (100%) when performed at the same temperature. After low-volume (2 ml extraction mixture) drug extraction from isolated leukocytes, the recoveries of DNR, DOL, and DOX from the cells were low, and they were inversely related to the cellularity of the sample, irrespective of the amount of drug in the cells. With high-volume extraction (5 ml extraction mixture) the recoveries were better (up to 95%), but they remained dependent on the cellularity. A correction factor accounting for these cellularity-related recoveries was applied to calculate the DNR and DOL contents of the leukocytes. Finally, using this information, plasma and cellular DNR and DOL levels were measured in seven patients with AML during their first course of remission induction therapy. The cellular DNR levels appeared to vary over a broad range and did not correlate with plasma pharmacokinetics.

Postirradiation thymocyte regeneration after bone marrow transplantation. III. Intrathymic differentiation and development of thymocyte subpopulations.

Fluorescence distributions of thymocytes stained for Thy-1 as well as size measurements were used to discriminate between thymocyte subpopulations during regeneration of the thymus after irradiation and bone marrow transplantation. Subpopulations with "low" and "high" Thy-1 density of donor- and recipient-derived progeny were quantitated. They were continuously present in the thymus and developed simultaneously but at different rates of growth. A similar developmental pattern was observed for donor- and host-derived "high" Thy-1+ cells, whereas "low" Thy-1+ cells of donor and recipient origin showed markedly different growth patterns. This indicated that development of the two subpopulations took place independently. During early regeneration donor-derived "low" and "high" Thy-1+ cells contain a high proportion of large cells, indicating the presence of cycling cells in both subpopulations.