An outbreak of Burkholderia cenocepacia bacteremia in immunocompromised oncology patients.

BACKGROUND: Burkholderia cepacia is a common environmental bacterium that is resistant to disinfectants, and therefore is often encountered as a hospital-acquired pathogen. We describe an outbreak of B. cenocepacia bacteremia among hospitalized oncology patients. METHODS: A matched case-control study and an extensive environmental investigation were conducted. Species were identified by RFLP of the amplified recA gene. DNA was fingerprinted by pulsed-field gel electrophoresis (PFGE). RESULTS: Between November 2005 and September 2006, B. cenocepacia bacteremia developed in 17 patients with underlying malignancy of whom 14 had tunneled central venous catheters. All patients had fever and chills which subsided following removal of the central catheter and administration of ceftazidime. Extensive epidemiological investigation could not find a common source for the outbreak. Patients were hospitalized in three different buildings with different health care personnel. Medications were prepared in different sites by different personnel. A multivariate analysis demonstrated that the independent risk factors for developing nosocomial B. cenocepacia bacteremia were hospitalization at the center for long-term support (OR 28.8; 95% CI 1.83-453.4) and reduced use of antibiotics during the last month (OR 0.07; 95% CI 0.01-0.40). All isolates had identical antimicrobial susceptibility; PFGE indicated that a complex of closely related strains was involved in the outbreak. All isolates were identified as B. cenocepacia, known to infect cystic fibrosis patients. Strict infection control measures terminated the outbreak. CONCLUSIONS: B. cenocepacia is an emerging nosocomial pathogen among oncology patients.

The Schneiderian membrane contains osteoprogenitor cells: in vivo and in vitro study.

Recent studies successfully demonstrated induction of new bone formation in the maxillary sinus by mucosal membrane lifting without the use of any graft material. The aim of this work was to test the osteogenic potential of human maxillary sinus Schneiderian membrane (hMSSM) using both in vitro and in vivo assays. Samples of hMSSM were used for establishment of cell cultures and for histological studies. Flow cytometry analysis was performed on P(0), P(1), and P(2) cultures using established mesenchymal progenitor cell markers (CD 105, CD 146, CD 71, CD 73, CD 166), and the ability of hMSSM cells to undergo osteogenic differentiation in culture was analyzed using relevant in vitro assays. Results showed that hMSSM cells could be induced to express alkaline phosphatase, bone morphogenic protein-2, osteopontin, osteonectin, and osteocalcin and to mineralize their extracellular matrix. Inherent osteogenic potential of hMSSM-derived cells was further proven by in vivo experiments, which demonstrated the formation of histology-proven bone at ectopic sites following transplantation of hMSSM-derived cells in conjunction with an osteoconductive scaffold. This study provides the biological background for understanding the observed clinical phenomena in sinus lifting. Our results show that a genuine osteogenic potential is associated with the hMSSM and can contribute to development of successful sinus augmentation techniques.

Human/mouse radiation chimera generated from PBMC of B chronic lymphocytic leukemia patients with autoimmune hemolytic anemia produce anti-human red cell antibodies.

Previous studies performed in our laboratory have shown that B-CLL cells are involved in the production of anti-red cell auto-antibodies, providing a possible mechanism for the autoimmune hemolytic anemia occurring during the course of B-CLL. In order to confirm this hypothesis, we attempted to transfer human B-CLL with AIHA to immunodeficient mice. Peripheral blood mononuclear cells (PBMC) from 11 B-CLL patients suffering from AIHA were transplanted into the peritoneal cavity of lethally irradiated Balb/c mice reconstituted with SCID bone marrow. Chimeric mice generated from PBMC of these patients (in stage III-IV of the disease) exhibited an engraftment profile with dominance of tumor cells and minuscule levels of T cells. Eighty-five percent of the chimeric mice generated from 10 out of the 11 B-CLL patients with Coombs'-positive AIHA, produced human Ig with anti-human red cell specificity as detected by indirect anti-globulin test. In addition, anti-red cell auto-antibodies were produced in 36% of chimeric mice generated from PBMC of Coombs'-negative B-CLL. In contrast, control experiments in which splenic cells from idiopathic AIHA or PBMC from normal donors were transplanted, failed to produce anti-RBC. This in vivo model further supports the relationship between the B cell expansion and the autoimmune hemolytic process.

Angiotensin converting enzyme inhibitors: 1,5-benzothiazepine derivatives.

The synthesis of chiral 1,5-benzothiazepines 2a-c, 14a-c, 15c, and 16a prepared from cysteine is described. In vitro inhibition of angiotensin converting enzyme (ACE) is reported for each compound. Compound 2c was the most potent in vitro having an IC50 of 2.95 nM. The ester of 2c, i.e. 14c, was found to inhibit the AI pressor response by 75% at a dose of 0.05 mg/kg iv and by 39% at 1.0 mg/kg po. Additionally, 14c lowered blood pressure in the spontaneous hypertensive rat (SHR) by 35 mmHg, at a dose of 10 mg/kg po.

hVISA and MRSA endocarditis: an 8-year experience in a tertiary care centre.

It is not clear if patients with heterogeneous intermediate resistance to vancomycin (hVISA) infectious endocarditis (IE) differ from methicillin-resistant S. aureus (MRSA) IE patients. All cases of hVISA and MRSA IE diagnosed at the Sheba Medical Centre from 2003 to 2010 were included. Isolates were screened prospectively for hVISA. Medical records were reviewed. The t-test, chi-square test, Fisher exact test and Kaplan Meier analysis were used. Fourteen hVISA IE and 32 MRSA IE were identified. The mean age was 76 years, mean Charlson score was 4.5 and 24% of patients had prosthetic valves. Pacemakers and implantable cardioverter-defibrillators (P/ICDs) were more common in the hVISA group (50% vs. 22%, p 0.05). P/ICDs IE occurred in 29% of hVISA patients vs. 6.3% of MRSA patients (p 0.06). hVISA patients had more positive blood cultures (eight vs. five, p 0.007) and a trend toward longer bacteraemia (15 vs. 7.5 days, p 0.08). Vancomycin minimal inhibitory concentrations (MICs) were similar in the two groups (1.5 µg/mL vs. 1.1 µg/mL, p 0.11). The MIC to daptomycin was higher in hVISA (0.75 µg/mL vs. 0.32 µg/mL, p 0.049). MRSA patients received vancomycin. hVISA patients were switched to other antibiotics. Cardiac surgery and/or P/ICD extraction was performed more commonly in hVISA patients (50% vs. 16%, p 0.027). Mortality was high in both groups (57-66%). The median time to death was 39 days in the hVISA group and 19 days in the MRSA group (p 0.3). hVISA IE is associated with P/ICDs. Both hVISA and MRSA are associated with high mortality. Low rates of surgical intervention and P/ICD extraction reflect the high co-morbidity of patients. Caution should be employed in the empirical use of daptomycin in hVISA patients.

Outcome of carbapenem resistant Klebsiella pneumoniae bloodstream infections.

The aim of this study was to evaluate the impact of carbapenem-resistant K. pneumoniae bloodstream infections on mortality. During the study period 42, 68 and 120 patients were identified with carbapenem-resistant, extended-spectrum ß-lactamase producers (ESBL) and susceptible K. pneumoniae bloodstream infections, respectively. Patients with carbapenem-resistant K. pneumoniae had higher rates of prior antimicrobial exposure, other nosocomial infections, and use of invasive devices. Infection-related mortality was 48% for carbapenem-resistant, 22% for ESBL producers and 17% for susceptible K. pneumoniae. Independent risk factors for infection-related mortality were Pitt bacteraemia score, Charlson score and carbapenem resistance.

Effects of FGF2 and FGF9 on osteogenic differentiation of bone marrow-derived progenitors.

Bone repair is a major concern in reconstructive surgery. Transplants containing osteogenically committed mesenchymal stem cells (MSCs) provide an alternative source to the currently used autologous bone transplants which have limited supply and require additional surgery to the patient. A major drawback, however is the lack of a critical mass of cells needed for successful transplantation. The purpose of the present study was to test the effects of FGF2 and FGF9 on expansion and differentiation of MSCs in order to establish an optimal culture protocol resulting in sufficient committed osteogenic cells required for successful in vivo transplantation. Bone marrow-derived MSCs cultured in αMEM medium supplemented with osteogenic supplements for up to three passages (control medium), were additionally treated with FGF2 and FGF9 in various combinations. Cultures were evaluated for viability, calcium deposition and in vivo osteogenic capacity by testing subcutaneous transplants in nude mice. FGF2 had a positive effect on the proliferative capacity of cultured MSCs compared to FGF9 and control medium treated cultures. Cultures treated with FGF2 followed by FGF9 showed an increased amount of extracted Alizarin red indicating greater osteogenic differentiation. Moreover, the osteogenic capacity of cultured cells transplanted in immunodeficient mice revealed that cells that were subjected to treatment with FGF2 in the first two passages and subsequently to FGF9 in the last passage only, were more successful in forming new bone. It is concluded that the protocol using FGF2 prior to FGF9 is beneficial to cell expansion and commitment, resulting in higher in vivo bone formation for successful bone tissue engineering.

A tissue-like construct of human bone marrow MSCs composite scaffold support in vivo ectopic bone formation.

Biocompatible and osteoconductive cell-scaffold constructs comprise the first and most important step towards successful in vivo bone repair. This study reports on a new cell-scaffold construct composed of gelatin-based hydrogel and ceramic (CaCO(3)/beta-TCP) particles loaded with human MSCs producing a tissue-like construct applied as a transplant for in vivo bone formation. Bone marrow-derived human MSCs were cultured in osteogenic induction medium. 5 x 10(5) (P(2)) cells were loaded on a mixture of hydrogel microspheres and ceramic particles, cultured in a rotating dynamic culture for up to 3 weeks. Both hydrogel microspheres and ceramic particles coalesced together to form a tissue-like construct, shown by histology to contain elongated spindle-like cells forming the new tissue between the individual particles. Cell proliferation and cell viability were confirmed by Alamar blue assay and by staining with CFDA, respectively. FACS analysis conducted before loading the cells, and after formation of the construct, revealed that the profile of cell surface markers remained unchanged throughout the dynamic culture. The osteogenic potential of the cells composing the tissue-like construct was further validated by subcutaneous transplants in athymic nude mice. After 8 weeks a substantial amount of new bone formation was observed in the cell-construct transplants, whereas no bone formation was observed in transplants containing no cells. This new cell construct provides a system for in vivo bone transplants. It can be tailored for a specific size and shape as needed for various transplant sites and for all aspects of regenerative medicine and biomaterial science.

The innate osteogenic potential of the maxillary sinus (Schneiderian) membrane: an ectopic tissue transplant model simulating sinus lifting.

Maxillary sinus membrane lifting is a common procedure aimed at increasing the volume of the maxillary sinus osseous floor prior to inserting dental implants. Clinical observations of bone formation in sinus lifting procedures without grafting bone substitutes were observed, but the biological nature of bone regeneration in sinus lifting procedures is unclear. This study tested whether this osteogenic activity relies on inherent osteogenic capacity residing in the sinus membrane by simulating the in vivo clinical condition of sinus lifting in an animal model. Maxillary sinus membrane cells were cultured in alpha-MEM medium containing osteogenic supplements (ascorbic acid, dexamethasone). Cultured cells revealed alkaline phosphatase activity and mRNA expression of osteogenic markers (alkaline phosphatase, bone sialoprotein, osteocalcin and osteonectin) verifying the osteogenic potential of the cells. Fresh tissue samples demonstrated positive alkaline phosphatase enzyme activity situated along the membrane-bone interface periosteum-like layer. To simulate the in vivo clinical conditions, the membranes were folded to form a pocket-like structure and were transplanted subcutaneously in immunodeficient mice for 8 weeks. New bone formation was observed in the transplants indicating the innate osteogenic potential within the maxillary Schneiderian sinus membrane and its possible contribution to bone regeneration in sinus lifting procedures.

Exposure to pro-inflammatory cytokines upregulates MMP-9 synthesis by mesenchymal stem cells-derived osteoprogenitors.

An intimate interplay exists between the bone and the immune system, which has been recently termed osteoimmunology. The activity of immune cells affects the intrinsic balance of bone mineralization and resorption carried out by the opposing actions of osteoblasts and osteoclasts. The aim of this study was to determine the possible interaction between inflammatory-induced conditions and matrix metalloproteinases-2,-9 (MMP-2,-9) synthesis and secretion by bone marrow-derived osteoprogenitor cells during advanced stages of osteogenesis. Rat bone marrow-derived mesenchymal stem cells (MSCs) were cultured in the presence of osteogenic supplements in order to direct the cells towards the osteogenic differentiation lineage. At the late stages of osteogenesis, assessed by histochemistry, immunohistochemistry and RT-PCR, cultures were exposed to pro-inflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 alpha (IL-1 alpha). Biochemical, histochemical and molecular biology techniques were used to discern the influence of pro-inflammatory cytokines on MMP-2,-9 synthesis and secretion. Results indicated that MMP-9 synthesis and secretion were significantly induced after exposure to the cytokines (TNF-alpha, IL-1 alpha) treatment, while MMP-2 levels remained unchanged. These results indicate that in response to inflammatory processes, osteoblasts, in addition to osteoclasts, can also be involved and contribute to the process of active bone resorption by secretion and activation of MMPs.

Flow cytometric determination of basophils in whole blood with n-propyl Astra Blue iodide.

Within the past year, it has become apparent, in connection with its use on automatic flow cytometers, that the quality of commercially available Alcian Blue has significantly declined. A homologous series of alkylated (C1-C7) Astra Blue quaternary ammonium halides was prepared, characterized, and evaluated for the detection of basophils in whole blood. On the Technicon H6000 flow cytometer, the resolution of the basophil cluster from the main population of unstained white blood cells was found to depend on the chain length of the quaternizing alkyl group. Optimal basophil resolution was observed for the n-propyl derivative. Correlation of the new method vs Alcian Blue as the reference on the H6000 was expressed as follows: %Baso (Astra Blue) = 0.89% Baso (Alcian Blue) + 0.12% for 180 fresh whole blood samples. Within-run precision at a basophil differential count of 0.73% was characterized by SD = 0.11, identical to that obtained for Alcian Blue. Aqueous solutions of n-propyl Astra Blue iodide, in contrast to Alcian Blue, are thermally stable. Heating the reagent for 1 h at 100 degrees C did not alter solubility or cytochemical behavior. In contrast, parallel treatment of Alcian Blue yielded insoluble material by hydrolysis of the isothiouronium groups. The reagent for basophil detection comprises n-propyl Astra Blue iodide, lanthanum chloride, sodium chloride, Tween 20, and cetylpyridinium chloride. The Astra Blue derivatives were characterized by uv-vis, ir, percentage halide, paper chromatography, and 13C NMR.

Acute femoral artery thrombosis associated with functional protein C deficiency as the presenting manifestation of acute monocytic leukemia.

Acute leukemia may be linked to disturbances of the coagulation system, which are usually due to hyperviscosity because of disseminated intravascular coagulation or a high white cell count. We report a unique case of femoral artery thrombosis as the beginning symptom of acute monocytic leukemia (M5) associated with a low level of functional protein C. The patient had multiple recurrent episodes of thrombosis with low levels of functional protein C associated with the relapse of the leukemia. A review of the literature shows that this is an extremely rare phenomenon.

Sjögren's syndrome presenting as hypokalemic paralysis due to distal renal tubular acidosis.

A 57-year-old woman presented with a flaccid paralysis, muscle tenderness, and respiratory depression. Laboratory results demonstrated severe hypokalemia with hyperchloremic metabolic acidosis and abnormally acidified urine. The urinary anion gap was positive in the presence of acidemia, thus establishing the diagnosis of distal renal tubular acidosis (DRTA). The patient fully recovered after potassium and alkali replacement. Further investigation revealed Sjögren's syndrome as the underlying cause of DRTA.

Clinical characteristics of the West Nile fever outbreak, Israel, 2000.

West Nile (WN) virus is endemic in Israel. The last reported outbreak had occurred in 1981. From August to October 2000, a large-scale epidemic of WN fever occurred in Israel; 417 cases were confirmed, with 326 hospitalizations. The main clinical presentations were encephalitis (57.9%), febrile disease (24.4%), and meningitis (15.9%). Within the study group, 33 (14.1%) hospitalized patients died. Mortality was higher among patients >70 years (29.3%). On multivariate regressional analysis, independent predictors of death were age >70 years (odds ratio [OR] 7.7), change in level of consciousness (OR 9.0), and anemia (OR 2.7). In contrast to prior reports, WN fever appears to be a severe illness with high rate of central nervous system involvement and a particularly grim outcome in the elderly.

West Nile fever outbreak, Israel, 2000: epidemiologic aspects.

From August 1 to October 31, 2000, 417 cases of West Nile (WN) fever were serologically confirmed throughout Israel; 326 (78%) were hospitalized patients. Cases were distributed throughout the country; the highest incidence was in central Israel, the most populated part. Men and women were equally affected, and their mean age was 54+/-23.8 years (range 6 months to 95 years). Incidence per 1,000 population increased from 0.01 in the 1st decade of life to 0.87 in the 9th decade. There were 35 deaths (case-fatality rate 8.4%), all in patients >50 years of age. Age-specific case-fatality rate increased with age. Central nervous system involvement occurred in 170 (73%) of 233 hospitalized patients. The countrywide spread, number of hospitalizations, severity of the disease, and high death rate contrast with previously reported outbreaks in Israel.