Detection of cytogenetics abnormalities in chronic lymphocytic leukemia using FISH technique and their prognostic impact.

INTRODUCTION: Chronic lymphocytic leukemia (CLL) is a clonal lymphoproliferative disorder characterized by progressive accumulation of morphologically and immunophenotypically mature lymphocytes. Characterization of genomic aberrations may help to understand the pathogenesis of CLL and may give prognostic information independent from conventional clinical markers for a risk-adapted management of CLL patients. AIM: The aim of the present study is to determine the most common cytogenetics abnormalities between patients with CLL and its prognostic impact. PATIENTS AND METHODS: The present study was carried out on 20 adult patients presented with chronic lymphocytic leukemia. The patients were diagnosed on the basis of standard clinical (lymph node involvement and/or hepatosplenomegaly), hematological and immunophenotypic criteria for diagnosis of B-CLL. All cases were studied at the time of their diagnosis. FISH technique was successfully performed on PB samples using CLL LSI probes for ATM (11q22) / GLI (12q13) and 13q14/ p53 (17p13). RESULTS: For comparative statistical studies, the patients were divided into group I (patients with favorable outcome) and group II (patients with unfavorable outcome). All patients showed one or more cytogenetic abnormality with the prevalence of p53 in 16 patients out of 20 that perfectly correlated with the poor outcome of the patients. This is followed by deletion in the 13q14 and to a lesser extent deletion in ATM gene, but no one has exhibited amplification in the 12q13 locus. CONCLUSION: p53 deletion as a sole abnormality has a higher prognostic power than other cytogenetics abnormalities. The cytogenetics study using FISH panel for CLL patients in a complementary fashion to the other clinical and laboratory findings may overcome the pitfalls in the diagnosis and may also assess the assignment of therapeutic protocols for CLL patients according to the results of their cytogenetic analysis at the time of diagnosis. KEYWORDS: FISH, chronic lymphocytic leukemia, CLL, p53, cytogenetics, Egypt.

Total dissolved and particulate lead in the Western Harbor of Alexandria, a Mediterranean basin under stress.

The Western Harbor of Alexandria, the largest harbor of Egypt on the Mediterranean Sea, receives different internal and external untreated pollutants affecting dramatically its water, sediments and biota. Surface and bottom water samples were collected bimonthly from this harbor for studying vertical, regional and monthly distributions of total suspended matter (TSM), total dissolved lead (TDPb) and total particulate lead (TPPb). The high surface TSM values coincided with air-borne dust, organic aggregates, plankton productivity and discharged wastes. The high TSM averages in winter resulted mainly from turbulence of the water column by wind action. The highest averages of TDPb in the petroleum and coal basins in the Western Harbor reflect their effects on the water column, while the lowest averages of TDPb and TPPb were found at a location far away from pollution sources. For the water column, the maximum and minimum monthly averages of TPPb and TDPb respectively in June reflect the increase in the rate of lead uptake by phytoplankton abundant in summer, favored by temperature elevation. The annual mean concentration of TPPb was nearly double that of TDPb, indicating that lead was transported to the Western Harbor mainly in particulate form. This reflects the high TSM content in the harbor water onto which a considerable part of lead was adsorbed.

Migration of bacteria along synthetic polymeric fibers.

E. coli, S. epidermidis, and B. distasonis were observed to migrate readily along polymer fibers impressed upon the surface of nutrient agar. E. coli was also observed to migrate readily along polymer fibers embedded in brain-heart infusion agar. Within periods of about 24 h, migration distances of about several centimeters were observed. No migration was observed in control experiments conducted on or in the same media, but without fibers. Migration speed was greatest for E. coli and slowest for B. distasonis. Cell population density was found to decrease rapidly with distance from a source culture. Swimming motility or natural convection in liquid between fiber and gel appears to be improbable based on the expected dimensions of capillary-condensed liquid between fiber and gel.

Production of dextransucrase by Leuconostoc mesenteroides immobilized in calcium-alginate beads: I. Batch and fed-batch fermentations.

In batch fermentation Leuconostoc mesenteroides immobilized in calcium alginate beads produced a total dextransucrase activity equal to about 93% of that by free, suspended bacterial cells under comparable conditions in a bubble column reactor. Continuous sucrose feeding (5 g/L h) to the immobilized-cell culture in the airlift bioreactor increased production of enzymatic activity by about 107% compared with ordinary batch operation of this reactor. About 14% of the enzymatic activity produced by the immobilized cells appears as soluble activity in the cell-free broth compared with about 40% in case of free cells. In an airlift bioreactor, both the soluble and the intact (sorbed and entrapped) enzymatic activity produced by the immobilized bacterial cells was about 34% greater under automatic pH control, compared to that produced in a bubble column reactor with only manual pH control. During formation of dextran by intact enzyme within cells and beads, declines are observed in apparent enzymatic activity.

Production of dextransucrase and dextran by Leuconostoc mesenteroides immobilized in calcium-alginate beads: II. Semicontinuous fed-batch fermentations.

Cells of Leuconostoc mesenteroides immobilized in calcium alginate beads were used to produce dextransucrase (DS) in three sequential cycles of semicontinuous fed-batch fermentations. Each cycle consisted of a fed-batch DS production period of 24 h followed by a batch dextran production period for another 24 h. Free, suspended cells were used in only one cycle of fed-batch DS production followed by a dextran production period. It was impractically tedious to separate and reuse free cells. Increasing sucrose feed rate from 5 to 10 g/L h led to increases of the total enzymatic activity by about 88% with immobilized cells and by about 100% with free cells. In DS fed-batch semicontinuous fermentation, total enzymatic activity produced by immobilized cells was 1.35 and 1.56 times greater than that produced by free cells with respective sucrose feeding rates of 10 and 5 g/L h. These increases in enzyme productivity with immobilized cells, however, required total overall operating times three times longer (three cycles) than with free cells (one cycle). Growing the microorganism at optimum conditions for DS production also increased the dextran yield and shortened the time of conversion of sucrose to dextran, regardless of whether the cells were free or immobilized. Moreover, during three cycles of semicontinuous operation (144 h) immobilized cells produced more than three times as much dextran as free cells during one cycle (24 h).

Effect of beta-cyclodextrin on production of L-phenylacetyl carbinol by immobilized cells of Saccharomyces cerevisiae.

The rate and extent of microbial transformation of higher concentrations of benzaldehyde substrate to L-phenylacetyl carbinol (L-PAC) by immobilized cells of Saccharomyces cerevisiae ATCC 834 was markedly stimulated by addition of different concentrations of beta-cyclodextrin (BCD) to the fermentation medium. With 0.5, 1.0, and 1.5% BCD in the fermentation medium and cumulative doses of benzaldehyde of 12 and 14 g/L, significantly higher yields of L-PAC were obtained, about one- to twofold that of the yields of the control experiments. The favorable effects of BCD were evident in spite of its presence in stoichiometric concentrations significantly lower than those of benzaldehyde. The presence of BCD also appeared to stimulate microbial growth slightly. Enhanced cellular activity was reflected by faster D-glucose consumption and faster benzaldehyde utilization in the presence of BCD.

Production of L-phenylacetyl carbinol by immobilized yeast cells: I. Batch fermentation.

Immobilization of Saccharomyces cerevisiae ATCC 834 within alginate beads enhances microbiological conversion of benzaldehyde to L-phenylacetyl carbinol (L-PAC), a precursor employed for synthesis of L-ephedrine. Yields of 90% L-PAC on benzaldehyde (initially 0.6% in medium) were obtained with immobilized cells, in contrast to about 10% with free cells which tend to form pellets in the presence of benzaldehyde. The predominant favorable action of immobilization appears to be a reduction in the toxic or inhibitory effects of benzaldehyde. With an initial benzaldehyde concentration of about 0.6% in the medium the optimum cell mass concentration was observed to be about 28 g cell mass (immobilized) per liter of medium.

Production of L-phenylacetyl carbinol by immobilized yeast cells: II. Semicontinuous fermentation.

The cyclic, semicontinuous production of L-phenylacetyl carbinol (L-PAC) from a benzaldehyde substrate by Saccharomyces cerevisiae ATCC 834 immobilized in calcium alginate beads was substantially enhanced to about 4.5 g/L in a second cycle by reactivation in fresh medium for 24 h, following an earlier 24-h period of production from substrate. Intermittent feeding of benzaldehyde was employed (four doses in 3 h). In subsequent similar cycles, however, the production returned to that produced in the first cycle, viz. L-PAC concentration of 2-3 g/L in the medium. Production of L-PAC was also increased by adaptation of the cells over 200 h of exposure to the benzaldehyde substrate (compared to wild-type cells) and by continuous (as compared to intermittent) feeding of the substrate. A liter as great as 10 g/L was obtained with wild-type cells by continuous feeding of benzaldehyde over 6 h. Immobilization not only protected the cells from toxic effects of substrate but also permitted them to be used during 7 cycles of semicontinuous operation over more than 200 h.

Comparative study of production of dextransucrase and dextran by cells of Leuconostoc mesenteroides immobilized on Celite and in calcium alginate beads.

In fed-batch fermentation, cells of L. mesenteroides immobilized on three types of Celite were used to produce dextransucrase (DS) followed by production of dextran. A layer of calcium alginate on the porous Celite R630 particles improved their mechanical stability, increased the amount of soluble DS produced and decreased the cell leakage from the highly porous support. Enzyme production with the immobilized cell cultures was significantly affected by both pore and particle size. Immobilized cultures using Celite R648 (average particle radius of 200 microm and pore size of 0.14 microm) produced the highest total enzymatic activity, followed by Celite R633, alginate-coated Celite R630, Celite R630, and then calcium alginate beads. Culture of free cells produced about 18% more total enzymatic activity than immobilized cells in calcium alginate beads, but about 64% less than immobilized cells on Celite R630. It is expected that larger amounts of enzymatic activity than measured are immobilized inside the alginate-coated Celite R630 and calcium alginate beads due to the mass transfer limitation conferred by the dextran product formed therein. The dextran yield from conversion of sucrose to dextran and fructose with all such enzyme-enriched, immobilized-cell cultures was higher than that obtained from free-cell culture under similar conditions.