OBJECTIVES:Superficial swab sampling of American tegumentary leishmaniasis (ATL) lesions shows higher amounts of Leishmania than those from biopsy. Subcutaneous involvement is also important in ATL, but parasite quantification according to lesion depth has not been evaluated. We aim to present the best depth at which sampling should be performed for molecular exams of ATL. METHODS:Patients with a clinical presentation compatible with ATL were allocated to ATL and control groups. Qualitative and quantitative qPCR assays were performed using SYBR Green and primers amplifying the kDNA minicircle of Leishmania spp. in different skin layers, including the epidermis, the superior dermis, the inferior dermis, and the hypodermis. RESULTS:Fifty-nine patients were included in this study, including 40 who had been diagnosed with ATL and 19 controls. The number of parasites was greater in samples of the epidermis and superior dermis (159.1 × 106, range 4.0-781.7, and 75.4 × 106, range 8.0-244.5, mean Leishmania parasite equivalents per μg of tissue DNA, respectively) than those in samples of the inferior dermis and hypodermis (54.6, range 8.0-256.6, and 16.8 × 106, range 8.0-24.1, mean Leishmania parasite equivalents per μg of tissue DNA, respectively). The best diagnostic accuracy was achieved in the superior dermis (77.9%) and was significantly greater than that in the hypodermis (63.3%; p 0.039). CONCLUSIONS:We conclude that superficial sampling can retrieve a greater quantity of parasites. Future studies of the role of transepidermal elimination as a mechanism of host defence in ATL must be performed as there is a considerable quantity of Leishmania kDNA in the epidermis.

SCF has been shown to synergize with G-CSF to mobilize CD34(+) PBPCs. In this study we report results from this combination after a phase II trial of 32 patients with malignant lymphoma randomized to receive recombinant methionyl human SCF (ancestim, r-metHuSCF) in combination with recombinant methionyl human G-CSF (filgrastim, r-metHuG-CSF) (experimental arm A) or routine chemotherapy plus filgrastim (conventional arm B). The primary objective was to evaluate the side effects and toxicity during priming and mobilization. The secondary objectives were efficacy by the level of blood-circulating PBPCs, the number of harvest days and the time to three-lineage engraftment after autografting. First, during priming 5 patients had 8 serious events, 4 in each arm. A summary of all adverse events revealed 30 (94%) patients suffering from 132 events of all grading. Second, neutropenia and thrombocytopenia was documented in arm B. Third, 9/14 (64%) patients in arm A reached the target of 5 million CD34(+) cells/kg body weight (bw) compared with 13/15 (87%) in arm B. The results represent the first randomized trial of growth factor plus chemotherapy priming and indicate that a formal phase III trial very unlikely may challenge chemotherapy plus r-metHuG-CSF priming in candidates for high-dose therapy.

Diagnosing candidaemia remains difficult despite the development of new diagnostics. We report a direct comparison of three different blood-culture systems and four indirect tests. One hundred and fourteen episodes either with haematological disease and fever despite antibacterials, or with documented invasive candidiasis, were enrolled prospectively. Clinical, para-clinical information and surveillance cultures were obtained. Blood culture was performed using conventional blood-culture bottles, mycosis bottles, and the Isolator 10 lysis centrifugation system. Serum D-arabinitol/L-arabinitol (DA/LA) ratios were determined by gas chromatography mass spectrometry. Antigen, mannan-antigen (Ag) and anti-mannan antibody (Ab) were detected by CandTec, Platelia Candida Ag ELISA and Candida AB/AC/AK kits, respectively. Episodes were classified as proven (n = 24), probable (n = 14), possible (n = 52) or unlikely (n = 24) invasive candidiasis. Candidaemia involved C. albicans (17), C. albicans + C. glabrata (3), C. tropicalis (1) and yeast (1). Mycosis bottles yielded two additional positives and the conventional blood culture yielded one positive not identified by other blood-culture methods. Considering proven and unlikely episodes, respectively, sensitivity and specificity were as follows: mannan-Ag and/or anti-mannan Ab: 83.3%, 78.3%; DA/LA ratio: 41.7%, 86.4%; and CandTec Candida Ag: 66.6%, 70.8%. Lowering the cut-off values to mannan-Ag 0.10 ng/mL and anti-mannan Ab 4 AU/mL, the values were: 100%, 73.9%. Applying the DA/LA ratio to only patients with haematological neutropenia the values were: 75%, 90.5%. Fungal blood culture allowed slightly improved detection of candidaemia. The best indirect test performance was obtained from combined mannan-Ag and anti-mannan Ab detection, especially with lower cut-offs. DA/LA ratio appears to be useful in the context of haematological neutropenia.

PURPOSE:YKL-40 is secreted by cancer cells, macrophages, and neutrophils. It may be a growth or differentiation factor, play a role in angiogenesis, or protect against apoptosis. High serum YKL-40 is associated with poor prognosis in solid carcinomas. The aim was to examine serum YKL-40 in patients with acute myeloid leukemia (AML).EXPERIMENTAL DESIGN:YKL-40 was measured by ELISA in serum from 77 patients recently diagnosed with AML before and during the first month of chemotherapy.RESULTS:Forty (52%) of the AML patients had elevated serum YKL-40 (compared with age-matched healthy subjects) and their survival was shorter than in patients with normal serum YKL-40 (median, 128 days; interquartile range, 18-629 days versus 386 days; interquartile range, 180-901; P=0.018 Mann-Whitney test). Univariate analysis of serum YKL-40 (logarithmically transformed and treated as a continuous covariate) showed significant association with survival within the first month after start of chemotherapy [hazard ratio (HR), 1.7; 95% confidence interval (CI), 1.2-2.4; P=0.002], first 12 months (HR, 1.6; 95% CI, 1.2-2.0; P=0.0002), and overall survival (HR, 1.3; 95% CI, 1.1-1.6; P=0.003). Multivariate Cox analysis showed that serum YKL-40 was an independent prognostic variable for survival (first month: HR, 1.7; P=0.011; 12 months: HR, 1.6; P=0.0002; overall survival: HR, 1.4; P=0.002). High serum YKL-40 at start of chemotherapy was a risk factor for pneumonia within the first month, and serum YKL-40 increased (P=0.002) at time of pneumonia and was unchanged in patients without infections.CONCLUSIONS:Serum YKL-40 is a prognostic biomarker of survival in AML patients. Its role in AML and infections needs to be determined.

Haemorrhage is often responsible for the lethal course of acute myeloid leukaemia (AML). Previously, multiple platelet function defects were identified by flow cytometric analysis of platelet activation markers in AML. The role of flow cytometric analysis of platelet function in characterization of prognostic markers of haemorrhage in AML patients has not been well elucidated. The objective of this prospective study was to analyse platelet function in 50 AML patients at diagnosis and to compare results with clinical bleeding score, graded by common toxicity criteria. Platelet activation markers CD62P, CD42b, CD63 and PAC-1 were analysed following in vitro activation by thrombin receptor activating peptide. The following plasma haemostasis parameters were measured: soluble P-selectin, activated partial thromboplastin time, thrombin time, prothrombin time, D-dimer, fibrinogen, and von Willebrand factor antigen. In a multivariate analysis, P-selectin (CD62P) <36 molecules of equivalent soluble fluorochrome x 10(3) (P < 0.0015) and platelet count <40 x 10(9)/l (P = 0.01) were significant predictors of haemorrhage at diagnosis. Haemorrhage at diagnosis predicted grade 3-4 haemorrhage in the first 28 d following diagnosis (P = 0.018). The presented results indicate that low P-selectin is a prognostic marker of haemorrhage in AML.

The most important prognostic factor in current clinical management of acute myeloid leukemia (AML) is the leukemic karyotype, identifying patients with relatively favorable, intermediate and adverse prognosis. Unfortunately, the majority of patients are assigned to the intermediate risk group although they represent a broad spectrum regarding prognosis. While molecular mapping may diminish this problem in the future, an approach to prognostic stratification of patients within the intermediate risk group is urgently needed in order to develop risk-adapted treatment strategies and evaluate the efficacy of new treatments. The aim of this retrospective study was to evaluate the prognostic impact of diagnostic multiparametric flow cytometry (MFC) in the cytogenetic intermediate risk group of AML patients. Diagnostic samples were obtained from 162 AML patients (median age 66 years, range 0–88; cytogenetic risk group favorable/intermediate/adverse/unknown 11/91/17/43; bone marrrow/blood 141/21). Four-color flow cytometry and a panel of 35 monoclonal antibodies was used to characterize the immunophenotype of AML blast cells, defined by side scatter and CD45 expression. For each antigen in the panel, mean fluorescence intensity (MFI) of the blast cell population was recorded. For each antigen, the expression pattern was characterized as homogeneous or heterogeneous. Heterogeneous antigen expression was defined as the presence of two or more subpopulations, within the blast cell gate, expressing the antigen in question with distinctly different intensities. Flow cytometry data, biological and clinical variables for the cytogenetic intermediate risk group and for patients with unknown cytogenetic risk were analysed by univariate Cox regression analysis and selected variables were analysed by multivariate Cox regression analysis (backward likelihood-ratio statistics, p<0.05 considered significant) with overall survival as outcome variable. Four conventional variables and four blast cell phenotype variables significantly predicting inferior survival were identified: FAB type other than 0/1 (p<0.03), presence of extramedullary disease (p<0.007), high white blood count (p<0.0005) and high age (p<0.0005); a heterogeneous CD117 expression (p<0.009), heterogeneous CD90 expression (p<0.03) as well as high MFI of CD56 expression (p<0.0005) and low MFI of CD58 expression (p<0.001) for CD45/SSC gated blast cells. The prognostic model identified two subsets of cytogenetic intermediate risk patients with a median overall survival of 0.1 and 1.5 years respectively. The subset of cytogenetic intermediate risk patients with high risk according to the prognostic model had an outcome inferior to that of the cytogenetically defined adverse risk group. This prognostic model was also valid for patients with unknown cytogenetic risk and for patients assigned to the favorable and adverse risk groups by cytogenetics. Informative MFC was applicable in 96% of the cases analysed. The prognostic model should be evaluated in a prospective clinical multicentre trial.

Abstract: Purpose: To estimate the clinical significance of low serum concentrations of mannose‐binding lectin (MBL) in patients with acute myeloid leukaemia (AML) during initial cancer chemotherapy. Patients and methods: 80 consecutive, newly diagnosed, and unselected AML patients (age 18–77 yr) undergoing remission induction chemotherapy. The patients were examined for 28 d. Main findings: Low levels of serum MBL (<1000 μg/L) were found in 16/80 patients at diagnosis. This frequency is similar to what is found in the general population. In the remaining 64 patients, MBL concentrations were significantly higher than in controls and showed only a slight rise during the period of antineoplastic chemotherapy with its associated infectious complications. Low levels of MBL did not affect overall survival or morbidity in terms of incidence or duration of fever, or occurrence of septicaemia or pneumonia. Long‐term survival was likewise independent of MBL concentration. Conclusion: MBL levels have no discernible influence on the occurrence or course of infections in AML patients during the initial hospitalisation. The predominant immunodeficiency during this phase is the profound granulocytopenia, which also compromises important effector functions of MBL. The finding in most AML patients of elevated MBL concentrations on admission is most likely because of the role of MBL as an acute phase reactant.

The clinical, histological, phenotypic and genotypic features of a lymphoblastoid natural killer (NK)-cell lymphoma presenting in the skin in a young caucasian woman are described. The disease behaved aggressively, but long-lasting remission was obtained by combination chemotherapy followed by autologous bone marrow transplantation. The blastoid cells were positive for terminal deoxynucleotidyl transferase, CD34, CD56 and CD4. Furthermore, the NK-cell receptor complex CD94/NKG2 was strongly expressed, as shown by examination with reverse transcription-polymerase chain reaction. The T-cell receptor (TCR)-gamma genes were in germline, and with the exception of CD4 all T-cell antigens were negative, including CD3, TCR-beta, TCR-delta, TIA-1, granzyme B and perforin. Epstein-Barr virus was negative, and no expression was seen of myeloid cell-associated markers. Molecular analysis showed no abnormalities of the CDKN2A (p16), CDKN2B (p15) or TNFRSF6 (Fas) genes. By contrast, a 34-bp deletion in exon 7 of the TP53 (p53) gene was detected. It is suggested that lymphoblastoid NK-cell lymphoma, which is a rare but distinctive disease, originates from NK cell precursors and may be associated with and possibly caused by alterations in the TP53 gene. Experience is too limited to warrant therapeutic suggestions. However, stem cell transplantation may be a useful option in younger patients.

Haemolytic anaemia caused by Clostridium perfringens is a rare complication in patients with neoplastic diseases. According to the literature, our patient seems to be the first patient with underlying malignancy (multiple myeloma) who has survived C. perfringens septicaemia complicated with acute haemolysis and acute anuria. It is concluded that treatment with penicillin should be considered in case of acute haemolytic anaemia, if the patient is febrile and no other obvious cause of the haemolytic condition can be found.

The purpose of this study was to evaluate the early infectious complications following autologous transplantation in haematoiogica! patients. Sixty-one patients who underwent either autologous bone marrow (BM; 28 patients) or peripheral blood stem cell (PBSC; 33 patients) transplantation for haematological malignancies were reviewed retrospectively. Engraftment happened significantly faster and the length of hospital stay was shorter in the PBSC group compared with the BM group. All patients in the study developed fever and all but two experienced temperatures ≥ 38.5°C. Overall, 57 patients had signs of oral mucositis, 23 with ulceration. Twenty patients had bacteraemia, 12 developed pneumonia, 6 systemic fungal infection. No major differences were found between the two groups in distribution or incidence of infections. This study indicates that the use of peripheral blood stem cells results in faster engraftment and shorter hospital stay, whereas the effect on the incidence of early infections seems to be unaffected.

PURPOSE A randomized, double-blind, placebo-controlled trial was performed to estimate the preventive effect of the antiherpetic drug acyclovir on fever, incidence of bacteremia, use of antibiotics, and presentation of infections in patients with acute myeloid leukemia (AML). PATIENTS AND METHODS Ninety herpes simplex virus (HSV)-seropositive patients aged 18 to 84 years were included. Forty-five patients received acyclovir (800 mg by mouth daily) and 45 placebo. The patients were examined daily for 28 days from the initiation of remission-induction chemotherapy. RESULTS Fever developed in all patients in both groups. Acyclovir prophylaxis postponed the development of an oral temperature > or = 38.0 degrees C by 3 days (95% confidence interval [CI], 1 to 4 days; P = .03) and the initiation of antibacterial treatment by 3 days (95% CI, 1 to 5 days; P = .008). The duration of fever, use of antibacterial treatment, incidence of bacteremia, and need for systemic antifungal therapy were not affected by acyclovir prophylaxis. At fever development, acyclovir prophylaxis affected the incidence and localization pattern of oral ulcers. Thus, in the acyclovir group, the number of nonfungal oral infections was reduced (relative risk, 0.45 [95% CI, 0.24 to 0.85]) and mainly located on the soft palate (relative risk, 2.49 [95% CI, 1.19 to 5.22]). CONCLUSION Acyclovir prophylaxis has an impact on fever development, but not on the duration of fever or the need for antibiotics. It does not reduce the incidence of bacteremia, but the presentation of acute oral infections is changed.

To investigate the oral coloniation of candidal pseudohyphae, a salivary smear method was tested against the use of mucosal smears in 10 patients with acute myeloid leukemia. The salivary smears showed to be reproducible and more sensitive than mucosal smears for the demonstration of Candida organisms. The salivary smear method was applied to another group of 10 patients with acute myeloid leukemia during a 28-day period. According to swab cultures, 8 patients were colonized with Candida spp. on day 0, whereas 2 patients were unclonized on day 0, and remained so. All colonized patients developed acute oral candidiasis. According to the salivary smear method, 5 patients were colonized with pseudohyphae as early as on day 0, whereas 3 patients became colonized with pseudohyphae within the first 7 days. Pseudohyphae were detected with a median of 4 days prior to the emergence of acute oral candidiasis, and the detection of acute oral candidiasis was associated with an increase in the quantity of pseudohyphoae. The study suggests that the demonstration of candidal pseudohyphae in salivary smears is an early indicator of subsequent development of acute oral candidiasis. Further studies are neded to clairfy whether early treatment based on this finding will reduce the occurrence of acute oral candidiasis and disseminated candidiasis in these patients.

Objectives-To evaluate (a) the prophylactic effect of the antiherpetic drug acyclovir on oral ulcers in patients with acute myeloid leukaemia receiving remission induction chemotherapy and thus (b), indirectly, the role of herpes simplex virus in the aetiology of these ulcers.Design-Randomised, double blind, placebo controlled trial.Subjects-74 herpes simplex virus seropositive patients aged 18-84. Thirty seven patients received acyclovir (800 mg by mouth daily) and 37 placebo. The patients were examined daily for 28 days.Main outcome measures-Occurrence of herpes labialis, intraoral ulcers, and acute necrotising ulcerative gingivitis.Results-The two populations were comparable in age, sex, type of antineoplastic treatment, and history of herpes labialis. Acute oral infections occurred in 25 of the acyclovir treated patients and 36 of the placebo treated patients (relative risk 0.69 (95% confidence interval 0.55 to 0.87)). This difference was due to a reduction in the incidence of herpes labialis (one case versus eight cases; relative risk 0.13 (0.02 to 0.95)), intraoral ulcers excluding the soft palate (one case versus 13 cases; relative risk 0.08 (0.01 to 0.56)), and acute necrotising ulcerative gingivitis (one case versus eight cases; relative risk 0.13 (0.02 to 0.95)). However, ulcers on the soft palate were diagnosed with similar frequency in the two groups. Isolation of herpes simplex virus type 1 in saliva was reduced from 15 cases in the placebo group to one case in the acyclovir group (relative risk 0.07 (0.01 to 0.48)).Conclusion-Intraoral ulcers excluding the soft palate are most often due to infection with herpes simplex virus, whereas ulcers on the soft palate have a non-herpetic aetiology. The findings suggest that acute necrotising ulcerative gingivitis may also be due to herpes simplex virus. Prophylaxis with acyclovir should be considered for patients with acute myeloid leukaemia during remission induction therapy.

Possible factors predisposing to the development of intraoral ulcers in immunocompromised patients with haematologic malignancies were investigated. Among 46 patients undergoing antineoplastic treatment, 18 developed an intraoral ulcer during the prospective study. Patients with or without ulcers were comparable with respect to underlying disease, presence of teeth, the qualitative composition of the aerobic and facultatively anaerobic oral microflora, herpes simplex virus (HSV) titer positivity, a past history of herpes labialis, leukocyte and thrombocyte counts, and duration of fever. Viral cultures from saliva revealed an association between the presence of HSV in saliva and the presence of intraoral ulcers. HSV was not isolated from the saliva of any patient without ulcers. Viral cultures from the ulcers revealed growth of HSV in 11 (61%) of the 18 patients with ulcers. HSV-positive ulcers were located more often on the alveolar process than elsewhere in the oral cavity. Ulcers on the lateral borders of the tongue and on the buccal mucosa were found only in dentate patients. Development of intraoral ulcers was not associated with an increase of the HSV titer in serum in any patient. It is concluded that some intraoral ulcers, especially when located on the alveolar process, are associated with and probably caused by HSV, while mechanical trauma during mastication seems likely to be involved in the pathogenesis of ulcers elsewhere in the oral cavity.

Abstract:To evaluate whether local oral humoral immunity changes during remission‐induction therapy of newly diagnosed acute myeloid leukaemia, 10 consecutive adult patients were investigated during 4 weeks. The concentrations of the oral secretory immunoglobulins A and M were increased during the first 7 days but, when corrected for changes in the salivary flow rate, the secretion rates of S‐IgA and S‐IgM remained unchanged, relative to the values before chemotherapy. In comparison, the concentrations of the serum immunoglobulins A, G and M were decreased, like the leukocyte and thrombocyte counts, during the first 14 d. It is concluded that the chemotherapeutic agents appear to act differently upon the local plasma cells associated with oral secretory glands and plasma cells in the bone marrow, and that the secretion rate of oral immunoglobulins remains constant during remission‐induction therapy.

To estimate clinical, pathogenic and serological aspects of acute oral candidiasis (AOC) during febril episodes in patients with haematologic malignancies, 23 consecutive patients who developed AOC within 7 days from start of fever were compared with 23 consecutive patients who did not develop AOC. The duration of fever and severe granulocytopenia (less than 0.5 x 10(9)/l) was significantly longer in patients with AOC than in patients without AOC, the median differences between the patients with and without AOC being 4 and 3 days, respectively. Development of AOC could not be correlated to a change in the qualitative composition of the oral microflora. The thrombocyte count was lower in patients with AOC on day 4, whereas no differences were found in leukocyte counts. The prevalences of Candida albicans agglutinin titres greater than or equal to 5 were similar in patients with (24%) and without AOC (33%), and in controls (29%). Seroconversion or a significant increase in the agglutinin titre occurred in 4 patients with AOC and long-lasting fever, who became afebrile after systemic antifungal therapy. It is concluded that AOC is associated with long-lasting fever and decreased bone marrow function as judged by low thrombocyte counts, but not related to specific bacteria in the oral cavity or to an increased occurrence of C. albicans antibodies in the serum.