Emerging resistant bacteria strains in bloodstream infections of acute leukaemia patients: results of a prospective study by the Rete Ematologica Lombarda (Rel).

Multiresistant bacterial infections are a potentially life-threatening condition in acute leukaemia (AL) patients. We aimed to better define the very recent epidemiology and outcome of bloodstream infections (BSIs) in a real-life setting. We prospectively collected all consecutive febrile/infectious episodes occurring in AL patients admitted to 9 haematology units. In 293 AL patients, 433 BSIs were diagnosed. Gram-positive (GP) bacteria were isolated in 44.8 % BSI and Gram-negative (GN) in 38.3 %, while polymicrobial aetiology- or fungi-related events were identified in 15.7 and 1.1 % of the cases, respectively. GP was observed more frequently in patients not in complete remission (p = 0.04), while GN during consolidation cycles (p = 0.003). Extended spectrum ß-lactamase-producing strains accounted for 23.2 % of enterobacteria. They were associated with previous antibiotic exposure, including fluoroquinolones prophylaxis (p = 0.01). Carbapenem-resistant (CR) strains occurred in 9 % of enterobacteria. Among Pseudomonas aeruginosa strains, 21.6 % were multiresistant. Overall 30-day mortality was 8.5 %. CR GN and multiresistant P. aeruginosa BSIs were independent predictors of death (p = 0.002), as well as relapsed/resistant AL (18.3 %; p = 0.0002) and the presence of pulmonary infiltrates (26.6 %; p < 0.001). Although GP still predominate over GN BSI, the percentage of antibiotic resistant GN strains is considerable in AL patients and it is associated with poor prognosis.

Effects of a new aminodiphosphonate (aminohydroxybutylidene diphosphonate) in patients with osteolytic lesions from metastases and myelomatosis. Comparison with dichloromethylene diphosphonate.

We have compared in an open trial the clinical and biochemical effects of a new aminodiphosphonate, aminohydroxybutylidene diphosphonate, with those of dichloromethylene diphosphonate, which has been proved effective. The patients presented extensive and symptomatic bone involvement from multiple myeloma, breast cancer, and other metastatic tumors. The treatment consisted of aminohydroxybutylidene diphosphonate, 2.5 mg/d intravenously for five days, or dichloromethylene diphosphonate, 300 mg/d intravenously for seven days, followed by 100 mg/d intramuscularly for ten days. Twelve patients treated with aminohydroxybutylidene diphosphonate and 16 patients treated with dichloromethylene diphosphonate were assessable and were followed up for one to six months. Therapy with aminohydroxybutylidene diphosphonate showed a quicker action in reducing bone pain and reduced significantly more the serum calcium level than did therapy with dichloromethylene diphosphonate. Aminohydroxybutylidene diphosphonate therapy also affected urinary calcium levels and hydroxyproline excretion more markedly than did dichloromethylene diphosphonate, although the differences are not statistically significant. However, the biochemical indexes rebounded more quickly in patients treated with aminohydroxybutylidene diphosphonate, indicating that the loading amount (only 12.5 mg) used in this preliminary study is insufficient to sustain a prolonged effect. The effectiveness and lack of side effects render aminohydroxybutylidene diphosphonate an attractive treatment for malignant bone resorption.

Development of simple equations for effective screening of spurious hemolysis in whole-blood specimens.

INTRODUCTION: We aimed to identify simple but reliable indices for effective screening of spurious hemolysis in whole-blood specimens. METHODS: Thirteen inpatient whole-blood samples were divided in two aliquots. The former was left untreated, whereas the latter was mechanically hemolyzed by forced aspiration with an insulin syringe. All aliquots were tested on Siemens Advia 2120 and Sysmex XE-2100. The hemolysis index (HI) was also assessed in centrifuged plasma. RESULTS: The mechanical hemolysis generated a 4-40% decrease in red blood cells (RBCs). A statistically significant decrease was observed for hematocrit (Ht) and mean corpuscular volume (MCV), whereas mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, and platelet count were increased. The values of hemoglobin (Hb) and white blood cells remained substantially unchanged. Two specific equations ([Ht/Hb] × âˆšMCV and [Ht/Hb] × 100) were developed. Both equations displayed an area under the curve of ≥0.99 for identifying spurious hemolysis, much greater than that of both RBC ghosts and immature platelet fraction. A highly significant correlation was also observed between results of these equations and percentage reduction in RBCs or HI increase. CONCLUSION: Provided that these results will be confirmed in further studies, these equations may provide a reliable means for screening spurious hemolysis in whole-blood samples.

Evaluation of the TiMo12Zr6Fe2 alloy for orthopaedic implants: in vitro biocompatibility study by using primary human fibroblasts and osteoblasts.

To reveal the biocompatibility of TiMo12Zr6Fe2 (TMZF), a new titanium alloy used since 1998 for orthopaedic prosthesis, we compared the behavior of primary human fibroblasts and osteoblasts grown on TMZF discs or on plastic tissue culture dishes, a widely used material specifically treated by the manufacturer to enhance cell growth. Proliferation, differentiation. RNA and collagen type I expression level of human cells were carried out. The analysis were performed over a period of 96 h. Fibroblasts behaved at the same way on the two different supports after 48 h, their number increased after 96 h when cells were grown on the alloy. Osteoblasts adhered and proliferated on the alloy discs as well as on plastic. RNA expression level was not affected. Interestingly, cell number at each time point was higher for fibroblasts than for osteoblasts. The RNA expression level was higher for the osteoblasts. Both cell types cultured on the alloy revealed an increase in the amount of type I collagen and a similar electrophoretic pattern was found for collagen produced by fibroblasts and osteoblasts grown on either supports. These results indicate good biocompatibility of the TMZF alloy, which allowed adhesion and proliferation of both the examined cell types and suggest that TMZF is a promising material for orthopaedic implants.

Clinical significance of serum pancreatic enzymes in the quiescent phase of chronic pancreatitis.

The most commonly used serum enzymes in pancreatic diseases are total amylase, pancreatic isoamylase, lipase and trypsin. To determine which of these enzymes is the most useful in the diagnosis of clinically quiescent chronic pancreatitis and which enzyme best reflects exocrine functional reserve, we studied 22 healthy control subjects, 44 patients with gastrointestinal, liver and biliary tract diseases, and 25 patients with chronic pancreatitis. On the basis of duodenal intubation, the latter were divided into two subgroups: one group of 13 patients with slight to moderate secretion deficiency and another of 12 patients with severe exocrine insufficiency. Of the enzymes studied, lipase, trypsin and pancreatic isoamylase are equally suitable for the evaluation of function in severe chronic pancreatitis, but not for the early diagnosis of the disease. Results for total amylase are not reliable so that its use in the study of chronic pancreatitis is not advisable.

Effect of Intralipid on some immunological parameters and leukocyte functions in patients with esophageal and gastric cancer.

This study has been undertaken to investigate if the intravenous (i.v.) infusion of fat emulsions may be associated with impairment of some immunological functions thus increasing the risk of septic complications. Fifteen malnourished patients with advanced gastric or esophageal cancer received for 2 weeks preoperatively and 1 week after surgery an isocaloric and isonitrogenous TPN treatment with Intralipid (group A: n=8) or glucose alone (group B: n=7) as energy substrate. Cluster analysis of 11 nutritional parameters and some tests of the humoral and cellular immunity (IgG, IgM, C3c, Factor B; polymorphonuclear (PMN) cells, total lymphocytes, T and B lymphocyte counts; 'in vitro' PMN chemotaxis, adherence to nylon fibers, phagocytosis of latex particles) were sequentially determined. The incidence and severity of post-operative infections were investigated and a 'sepsis score' was calculated for each patient. Pre- and postoperative TPN were not associated with an improvement of the nutritional status. The humoral and cellular immune parameters showed the same behaviour in patients receiving Intralipid and in controls. The chemotactic activity of PMN cells was constantly normal, granulocyte adherence fluctuated below the normality range in controls, whereas phagocytosis of latex was similar in both groups. Post-operative infectious episodes were less severe in patients receiving Intralipid. Our results do not confirm that Intralipid adversely affects some aspects of the humoral and cellular immune response.

Efficiency of composite laboratory tests in the diagnosis of liver malignancies.

The utility of the markers CEA, beta-HCG, CA-50, alpha-fetoprotein (APF), ferritin, alkaline phosphatase (AP), its isoenzyme liver-1 (APL1), gamma-glutamyltransferase (gGT), its fast migrating isoenzyme (gGT1) and 5'nucleotidase (5'N) in differentiating liver malignancies and benign involvement was evaluated in the sera of 85 patients with hepatocellular carcinoma (HCC), 157 with chronic liver disease (CLD) and 91 with liver metastases (LM) derived from different tumors. The mean concentrations of all the parameters except CEA and GGT1 were significantly different in HCC and CLD, but a broad overlap existed in the two groups, so different cut-offs were considered to assess the positive and negative predictive values and test efficiency (Eff). The best results were observed considering AFP greater than 100 IU/m (Eff0.86), ferritin greater than 800 ng/ml (Eff0.69), CA-50 greater than 100 U/ml (Eff 0.63), beta-HCG greater than 10 mU/ml (Eff 0.61), AP greater than 300 IU/ml (Eff 0.66), the presence of APL1 (Eff 0.78), 5'N greater than 25 mU/ml (Eff 0.70), gGT greater than 100 mIU/ml (Eff 0.63). Among HCC patients 17% did not secrete AFP; in 26% the protein was less than 100 IU/ml and in 36% less than 400 IU/ml. Apart from AFP the most effective marker was APL1. At the above cut-offs more than three parameters were simultaneously positive in 71% of HCC and 9.9% of CLD. CEA, CA50, AFP were the only parameters that distinguished the HCC from the LM group; in the latter, APL1 was also a very sensitive marker (87%) for neoplastic involvement of the liver.

Tumour associated antigens CA 15.3 and CA 125 in ovarian cancer.

CA 125 and CA 15.3 antigens were determined by enzyme immunoassay in 78 patients with ovarian cancer for a total of 540 determinations. The antigens were also investigated in sera from 100 women with other gynaecological diseases, 82 lung cancer patients and in 39 pleural fluids of varying origin. CA 15.3 reference values were evaluated in 91 healthy women (cut-off: 25 U/ml). CA 15.3 sensitivity at diagnosis (60%) and for detecting relapse (44%) was lower than that of CA 125 (90% and 64.7%, respectively). However, CA 15.3 does not increase with aspecific mesothelial cell reaction and thus it is more specific than CA 125. Combined use of the markers during follow-up improves early detection of relapse (at least one of the two was positive in 79% of cases). Therefore both CA 15.3 and CA 125 should be routinely determined for the detection and monitoring of ovarian cancer.

Nutritional and immunological evaluations in cancer patients. Relationship to surgical infections.

Several reports provide evidence of the synergism between malnutrition and infection in hospitalized patients. In this study, preoperative complete nutritional assessment (NA) was performed to: 1) evaluate NA modifications in 21 controls with benign minor surgical diseases and in 71 surgical cancer patients; 2) determine the relative value of nutritional and immunological indicators in relation to the postoperative septic complications in cancer patients. The following parameters were used: percent weight loss/month, percent standard arm circumference, percent standard triceps skinfold; hematocrit, hemoglobin, serum proteins, albumin, iron, transferrin, ceruloplasmin, retinol binding protein (RBP); serum creatinine, urine creatinine, creatinine/height index, percent arm muscle circumference; peripheral lymphocytes, white blood cells, complement (C3c), skin tests. In the cancer group percent usual body weight, percent arm muscle circumference, hematocrit, hemoglobin, albumin, iron, ceruloplasmin, retinol binding protein, C3c and delayed hypersensitivity response (DHR) were significantly different from controls. Ceruloplasmin and DHR were the only tests significantly different in the cancer patients who developed postoperative infections. Duration of anesthesia and operative field contamination are other important causative factors which, if associated with malnutrition, may determine a higher susceptibility to infections in surgical cancer patients.

Evaluation of possible causes of delayed hypersensitivity impairment in cancer patients.

The separate roles of malnutrition, advanced age, and stage of tumor growth as causes of impairment of delayed hypersensitivity response (DHR) was studied in 111 patients with solid tumors and in 56 nonneoplastic control patients matched for age, anatomical site of disease, degree of illness, and nutritional status. Pretreatment DHR to recall antigens (tuberculin, Candida, streptokinase-streptodornase, trichophyton) and to dinitrochlorobenzene in cancer patients with 9% anergic, 43% hypoergic, and 48% normoergic; the distribution of DHR in controls was not significantly different. In cancer patients, the serum albumin level showed an inverse correlation with the stage of tumor (p less than 0.01) and a positive correlation with the DHR (p less than 0.001); the serum albumin level was also in the controls positively correlated with the DHR (p less than 0.01), indicating that malnutrition in neoplastic or benign disease may cause depression of DHR. In well-nourished controls, age was inversely correlated with DHR (p less than 0.05), showing that aging itself may be another relevant cause of depression of DHR. The results of this study indicate that DHR in patients with solid tumors is similar to the DHR of nonneoplastic patients if matched for age, sex, and nutritional status. DHR impairment in cancer patients appears to be caused mainly by aging and by malnutrition due to the advanced progression of cancer.

Peptichemio, vincristine, prednisone induction treatment in multiple myeloma.

Twenty-nine patients with multiple myeloma, 16 untreated and 13 relapsing after treatment with melphalan and/or cyclophosphamide, were treated with Peptichemio (PTC), vincristine (VCR) and prednisone (PRD). The treatment was well tolerated and produced rapid resolution of bone pain and improved performance status in symptomatic patients. Rapid and marked monoclonal component reduction (greater than 50 per cent) was noted in 50% of previously untreated and 31% of previously treated patients. The median survival of previously untreated patients was 27 months, 19 months for stage III patients. Four patients resistant to common alkylating agents responded to PTC-VCR-PRD treatment and achieved long remissions (from 18 to 38 months). The therapeutic results suggest that the present combination regimen may be indicated for induction treatment of multiple myeloma patients in an advanced stage of the disease, and for treatment of myeloma patients refractory or resistant to melphalan and/or cyclophosphamide.

Neer capsular shift surgery in the treatment of recurrent antero-inferior shoulder dislocations.

A total of 47 patients affected with recurrent multidirectional shoulder dislocation with a prevalent anteroinferior component were submitted to Neer inferior capsular shift surgery. Bankart lesion, present in 24 patients, was repaired prior to performing capsuloligamentous surgery. The patients included in the study presented with generalized ligamentous hyperlaxity in 20 cases, while hyperlaxity was localized in the pathological shoulder in the remaining cases, presumably related to the high number of dislocations. The choice of performing Neer inferior capsular shift surgery was initially based on the data and the modern conceptions that emerged in the literature. Successively, the good results obtained encouraged us to follow the path taken, and we are currently encouraged to advise this surgical procedure for the category of patients mentioned above.

Early molecular diagnosis of aspergillosis in a patient with acute myeloid leukaemia.

Diagnosis of invasive fungal infection remains challenging. Here we report a case of early diagnosis of invasive aspergillosis in a neutropenic patient affected by acute myeloid leukaemia, achieved through the detection of Aspergillus fumigatus species-specific ribonucleic acid sequences by a sensitive multiplex real-time polymerase chain reaction-based molecular assay. Thanks to the early diagnosis, targeted therapy was promptly established and the severe fungal infection controlled, allowing the patient to subsequently receive allogeneic hematopoietic stem cell transplantation from a haploidentical donor, her only curative option. Also in this instance, targeted secondary antifungal prophylaxis with voriconazole avoided any other fungal infection afterwards. This report suggests how the implementation of molecular assays in combination with routine diagnostic procedures, can improve microbiological diagnosis in sepsis, particularly in case of fungal infection, difficult to detect with standard microbiological culture methods.

Influence of in vitro hemolysis on nucleated red blood cells and reticulocyte counts.

INTRODUCTION: Nucleated red blood cells (NRBCs) and reticulocytes are early and important measures of red blood cells' (RBCs) turnover, but little is known on how spurious hemolysis may affect the reliability of these parameters. MATERIALS AND METHODS: Ten EDTA-anticoagulated samples were divided into three aliquots. The first was immediately tested, where-as the others (defined A and B) were mechanically hemolyzed by aspiration 5 and 10 times through a small-gauge needle. RBC, NRBC, and reticulocyte counts were performed on Sysmex XE-2100. RESULTS: An increasing amount of hemolysis was produced in hemolyzed aliquots A and B. The RBC and reticulocyte counts progressively decreased from the nonhemolyzed sample to hemolyzed aliquots 'A' and 'B'. The NRBC count increased in 3 of the 10 samples and decreased in the remaining seven. CONCLUSIONS: Hemolysis of venous blood samples may seriously jeopardize NRBC and reticulocyte counts.

Beta-2-microglobulin as a tumor marker in solid malignancies.

Beta-2-microglobulin (beta 2-MG) and CEA were measured in the sera of 186 cancer patients divided into two groups: at diagnosis (D) and at follow-up (F). Four groups of patients at diagnosis (D-I, D-II, D-III and D-IV according to TNM classification) and two at follow-up (in remission, F-RS, and in relapse, F-RP) were considered. All patients had normal serum creatinine and urea concentrations. beta 2-MG values in D-I were significantly (p less than 0.01) lower than those for D-II and D-III, while in D-IV they overlapped those of group D-I. No significant difference was observed between F-RS and F-RP patients. Patients with serum CEA concentration greater than 100 ng/ml revealed beta 2-MG values close to those of groups D-I and D-IV. In 10% of patients in stage IV or with CEA greater than 100 ng/ml beta 2-MG was lower than the mean value of the healthy population. From data beta 2-MG is probably produced by an aspecific reaction to the tumor and the decrease in advanced stages could express a decreased immunologic response. On the other hand, high serum beta 2-MG in the initial stages of the neoplasia may reflect an elevated cell turnover, while low beta 2-MG during the final stages may be due to a weak expression of the protein by highly undifferentiated cells.

Micropolyspora faeni antigens of three commercial extracts.

The protein composition of three commercial extracts of Micropolyspora faeni, produced in U.S.A., England and Italy has been evaluated by agarose gel electrophoresis. By crossed immunoelectrophoresis, 'tandem'-crossed immunoelectrophoresis and by a modification of this last technique, the antigenic composition and the common antigens of the extracts have been investigated. Hyperimmune rabbit serum and a pool of five human sera with precipitins to Micropolyspora faeni have been used as source of antibody. Different quantity and quality of protein content was observed in the available batches. Different antigenic composition was also observed, not directly related to the different proteins contained therein; three antigens were definitely common to all extracts and two of them represented the major antigens of each extract. Despite the total protein content, the major and common antigens were found in similar concentrations in all three products examined. Therefore, the discrepancies observed in the precipitin reactions using the three commercial Micropolyspora faeni extracts are due to differences in the minor antigen composition of the extracts.

Variations in alpha-2-HS glycoprotein level in neoplastic disease with and without involvement of bone.

Variations in the serum level of alpha-HS-glycoprotein may be observed in all pathological conditions which induce changes in bone turn-over, as well as in inflammatory and neoplastic diseases. This study includes 162 patients divided into four groups according to the TNM classification (tumour, lymph node metastasis). The first consisted of patients with neoplasms at TNM stages 1 and 2 with no bone metastases; the second of similar patients at TNM stages 3 and 4. The third group were patients with primary or secondary neoplasms of bone, and the fourth were patients with viral or bacterial diseases. The levels of alpha-2-HS-glycoprotein serum were determined for all the groups and these were compared with AAG (alpha-1-glycoprotein) and CEA (carcinoembryonic antigen). There were no significant differences in the levels of alpha-2-HS-glycoprotein for the first group as compared with normal controls, while in the other groups the differences were significant. The levels of alpha-2-HS-glycoprotein were diminished when the levels of CEA and AAG were both high, but increased when only one of these other parameters was high.

Detection of malignant pleural effusions by tumor marker evaluation.

Cytologic examination and determination of tumor markers (PHI, LDH, alpha-1-glycoprotein, alpha-2-HS-glycoprotein, beta 2-microglobulin, ferritin [corrected], sialic acid, IgE, fetoprotein, CEA, beta HCG and beta 1-SP-glycoprotein) were carried out in pleural fluid samples obtained from 70 patients with suspected neoplasia. Tumor markers were also determined in sera. The protein content of all pleural effusions was greater than or equal to 3 g/dl. Patients were grouped according to diagnosis as follows: (a) 42 with neoplastic diseases (7 mesotheliomas and 19 lung, 4 ovarian, 3 breast and 8 miscellaneous cancers), (b) 22 with benign inflammations and (c) 6 with congestive effusions. Of the parameters examined, only CEA and beta-HCG [corrected] gave information that the effusion was probably malignant. Using 6 ng/ml as cut-off for CEA and 10 mIU/ml for beta HCG, the sensitivity was 57.1% and 45.2%, respectively, specificity was 92.8% for both parameters and test efficiency 0.75 and 0.69, respectively. When CEA and beta HCG were considered together sensitivity increased to 73.8% and efficiency to 0.78. CEA and/or beta HCG were positive in the pleural effusions of 19 of the 20 malignant pleural effusions, all with a negative cytologic examination, which subsequently became positive in 8. Because of their high specificity, these two parameters are a useful tool and can be routinely measured to evaluate pleural effusions of dubious origin, even if CEA and beta HCG cannot, on [corrected] their own, define the primary malignancy.

Identification of specific plasma proteins determining the agarose gel electrophoresis by the immunosubtraction technique.

The immunosubtraction technique was used to identify the proteins in electrophoretically separated plasma. The technique consists of forcing the plasma through a layer of monospecific antibody purified by electrophoresis; the specific protein is thereby precipitated and is consequently absent from the subsequent electrophoretogram. This technique was successfully applied to the following proteins (in order of distance from the anode): prealbumin, albumin, alpha-lipoproteins, alpha 1-antitrypsin, Gc-globulins, alpha 2-macroglobulin, haptoglobin, fibronectin, transferrin, beta-lipoproteins, C3, fibrinogen. The method may be applied to the study of protein polymorphism.

Ring chromosome and latent centromeres.

A ring 12 chromosome was found in a male child with minor phenotypic alterations. No obvious loss of chromosome material was detected. Since there is no other case of a ring 12 in the literature, it was not possible to determine whether the phenotype was due to (invisible) terminal deletions or to karyotypic variation. In lymphocyte cultures 9% of the cells had either no or two rings, but the patient's RBC had normal activities of the enzymes lactate dehydrogenase B and peptidase B, whose loci are located on the proximal portions of 12p and 12q, respectively. Dicentric rings were found in 37 cells, and all had two active centromeres, in contrast with the relatively frequent finding of latent centromeres in translocated dicentric autosomes. Two latent centromeres were found in one tricentric "rod-shaped" open ring 12, probably derived from a tetracentric ring. It is postulated that latent centromeres are rare in ring chromosomes because, if consistent suppression of centromeres in excess of one took place at each duplication, rings with four, eight, or more centromeres would be formed rather frequently, which is not the case.