Lysis Products Research Articles

Influence of algal biomass on extracellular enzyme activity in river biofilms.

Relationships between biofilm structural components (algal and bacterial biomass) and the activities of some extracellular enzymes that contribute to the ability to degrade organic matter) were explored for six Atlantic and three Mediterranean streams and rivers. The biofilms in these fluvial systems accounted for a wide range of bacterial and algal biomass and colonized the most common benthic habitats. Ratio of bacteria/algae biomass was lower in Atlantic than in Mediterranean streams, but enzymatic activities (β-glucosidase, β-xylosidase, phosphatase) were in general greater in the Mediterranean stream biofilms. Climatic characteristics (especially temperature) may explain the differences in enzymatic activities between biofilms of similar structure but different flow regime. The ratio β-xylosidase: β-glucosidase was similar (around 0.5) for all streams and substrata considered, showing that there is a general higher utilization of cellobiosic than xylobiosic molecules in fluvial systems. In general, highly heterotrophic biofilms showed lower extracellular enzymatic activities than more autotrophic biofilms. Maximum enzymatic activity is achieved when the algal biomass is two- to threefold higher than the bacterial biomass. The relevance of algal biomass on the heterotrophic ability of biofilms may be related to the physical proximity between the two, but also to the high proportion of polymeric carbohydrates included in algal exudates and lysis products, whose use is enzyme-mediated.

Experimental study of the effects of Bacillus subtilis on gibbsite dissolution rates under near-neutral pH and nutrient-poor conditions

In order to examine the effects of Bacillus subtilis on rates and extents of gibbsite dissolution, we conducted batch-type dissolution experiments with viable, non-viable, and no bacteria. Dissolved Al and organic carbon were monitored at regular time intervals for approximately 10 days under near-neutral pH and nutrient-poor conditions that simulated the conditions under which natural fluid–mineral interactions occur. Viable bacteria not only significantly enhanced the gibbsite dissolution rate, but they also enhanced the extent of dissolution by a factor of 30 relative to the bacteria-free control experiments. The dissolved Al concentrations in the experimental solutions are correlated to the concentration of dissolved organic carbon, suggesting that organic molecules derived from the bacteria are responsible for the observed dissolution behavior. Non-viable bacteria also cause enhancement of the rate and extent of gibbsite dissolution relative to the controls, but the observed Al concentrations are five to seven times lower than those observed for the systems containing viable bacteria. This result implies that abiotic surface complexation between aqueous Al and the bacterial cell wall functional groups is not the cause of the dissolution enhancement. However, the dissolved organic carbon concentrations in the experiments containing non-viable bacteria are virtually identical to those observed for the systems containing viable bacteria. This similarity strongly suggests that the bacterially derived organic molecules from both types of bacteria are lysis products rather than metabolic exudates, and that the lysis products from intact bacteria are more capable of enhancing the rate and extent of gibbsite dissolution than those of the non-viable bacteria. These conclusions are supported by IC and GC-MS analyses of the two different types of dissolved organic carbon, and by results from dissolution experiments that were engineered to prevent bacterial lysis. The experimental results presented here are the first to indicate that bacterial lysis can significantly affect mineral dissolution in weathering environments, and that lysis products may be as important as bacterial exudates in controlling the rate and extent of mineral dissolution.

Influence of J series prostaglandins on apoptosis and tumorigenesis of breast cancer cells.

This study was undertaken to investigate the influence of the peroxisome proliferator-activated receptor gamma (PPARgamma) agonists on the proliferation, apoptosis and tumorigenesis of breast cancer cells. PPARgamma investigation has been largely restricted to adipose tissue, where it plays a key role in differentiation, but recent data reveal that PPARgamma is expressed in several transformed cells. However, the function of PPARgamma activation in neoplastic cells is unclear. Activation of PPARgamma with the known prostanoid agonist 15-deoxy-Delta12,14-prostaglandin J(2) (15dPGJ(2)) or the thiazolidinedione (TZD) agonist troglitazone (TGZ) attenuated cellular proliferation of the estrogen receptor-negative breast cancer cell line MDA-MB-231, as well as the estrogen receptor-positive breast cancer cell line MCF-7. This was marked by a decrease in total cell number and by an inhibition of cell cycle progression. Addition of 15dPGJ(2) was not associated with an increase in cellular differentiation, as has been seen in other neoplastic cells, but rather induction of cellular events associated with programmed cell death, apoptosis. Video time-lapse microscopy revealed that 15dPGJ(2) induced morphological changes associated with apoptosis, including cellular rounding, blebbing, the production of echinoid spikes, blistering and cell lysis. In contrast, TGZ caused only a modest induction of apoptosis. These results were verified by histochemistry using the specific DNA stain DAPI to observe nuclear condensation, a marker of apoptosis. Finally, a brief exposure of MDA-MB-231 cells to 15dPGJ(2) initiated an irreversible apoptotic pathway that inhibited the growth of tumors in a nude mouse model. These findings illustrate that induction of apoptosis may be the primary biological response resulting from PPARgamma activation in some breast cancer cells and further suggests a potential role for PPARgamma ligands for the treatment of breast cancer.

Rapid on/off cycling of cytokine production by virus-specific CD8+ T cells.

CD8-positive T cells protect the body against viral pathogens by two important mechanisms: production of antiviral cytokines and lysis of infected cells. Cytokine production can have both local and systemic consequences, whereas cytolytic activity is limited to infected cells that are in direct contact with T cells. Here we analyse activated CD8-positive T cells from mice infected with lymphocytic choriomeningitis virus and find that cytokines are not produced ex vivo in the absence of peptide stimulation, but that they are rapidly generated after T cells encounter viral peptides bound to the major histocompatibility complex. Remarkably, cytokine production ceases immediately upon dissociation of the T cells from their targets and resumes when antigenic contact is restored. In contrast to the 'on/off/on' cycling of cytokines, the pore-forming cytotoxic protein perforin is constitutively maintained. Our results indicate that there is differential expression of effector molecules according to whether the antiviral product is secreted (like cytokines) or stored inside the cell (like perforin). The ability to turn cytokines on and off while maintaining intracellular stores of perforin shows the versatility of the cellular immune response and provides a mechanism for maintaining effective immune surveillance while reducing systemic immunopathology.

Formation of Dimethyloligo-Sulfides in Lake Kinneret

Dimethyloligosulfides were recently identified as a primary source of mild malodorous emissions from Lake Kinneret, Israel. The seasonal odor episodes coincide with a bloom of Peridinium gatunense algae. The possibility that the dimethyloligosulfides are formed by bacterial degradation of Peridinium gatunense lysis products, under oxygen rich conditions was investigated. Several bacterial strains were isolated from the lake. Addition of Peridinium cells to the isolated bacteria cultures yielded dimethyldisulfide and dimethyltrisulfide. One of the bacteria strains, identified as Acinetobacter lwoffii, an obligatory aerobe was singled out for detailed investigation. Addition of Peridinium cells or methionine to the Acinetobacter culture yielded, under aerobic conditions dimethyldisulfide and dimethyltrisulfide. Cystein feed yielded only inorganic oligosulfides, which were converted to dimethylsulfides by addition of d3-methyliodide.

Formation of Dimethyloligosulfides in Lake Kinneret: Biogenic Formation of Inorganic Oligosulfide Intermediates under Oxic Conditions

The mechanism of formation of dimethyloligosulfides in Lake Kinneret was investigated by field and laboratory studies. The process was simulated under laboratory conditions using obligate aerobic and facultative bacteria that were isolated from Lake Kinneret and fed with different types of organo-sulfur nutrients. The lysis products of Peridinium gatunense--a dinoflagellate that dominates the phytoplankton population in Lake Kinneret during the winter-spring season--are the primary source of dimethyloligosulfides. Bacterial assimilation of the aged alga or algae lysis products yields inorganic oligosulfides, which are then methylated to form the dimethyloligosulfides. All the steps of this process are carried out under oxic conditions.

Breakdown and microbial uptake of marine viruses and other lysis products

Breakdown and microbial uptake of marine viruses and other lysis products

Breakdown and microbial uptake of marine viruses and other lysis products

Breakdown and microbial uptake of marine viruses and other lysis products

Formation of dimethyloligosulfides in lake kinneret

Dimethyloligosulfides were recently identified as a primary source of mild malodorous emissions from Lake Kinneret, Israel. The seasonal odor episodes coincide with a bloom of Peridinium gatunense algae. The possibility that the dimethyloligosulfides are formed by bacterial degradation of Peridinium gatunense lysis products, under oxygen rich conditions was investigated. Several bacterial strains were isolated from the lake. Addition of Peridinium cells to the isolated bacteria cultures yielded dimethyldisulfide and dimethyltrisulfide. One of the bacteria strains, identified as Acinetobacter lwoffii , an obligatory aerobe was singled out for detailed investigation. Addition of Peridinium cells or methionine to the Acinetobacter culture yielded, under aerobic conditions dimethyldisulfide and dimethyltrisulfide. Cystein feed yielded only inorganic oligosulfides, which were converted to dimethylsulfides by addition of d 3 -methyliodide.

Significance of viral lysis and flagellate grazing as factors controlling bacterioplankton production in a eutrophic lake.

The effects of viral lysis and heterotrophic nanoflagellate (HNF) grazing on bacterial mortality were estimated in a eutrophic lake (Lake Plusssee in northern Germany) which was separated by a steep temperature and oxygen gradient into a warm and oxic epilimnion and a cold and anoxic hypolimnion. Two transmission electron microscopy-based methods (whole-cell examination and thin sections) were used to determine the frequency of visibly infected cells, and a model was used to estimate bacterial mortality due to viral lysis. Examination of thin sections also showed that between 20.2 and 29.2% (average, 26.1%) of the bacterial cells were empty (ghosts) and thus could not contribute to viral production. The most important finding was that the mechanism for regulating bacterial production shifted with depth from grazing control in the epilimnion to control due to viral lysis in the hypolimnion. We estimated that in the epilimnion viral lysis accounted on average for 8.4 to 41.8% of the summed mortality (calculated by determining the sum of the mortalities due to lysis and grazing), compared to 51.3 to 91.0% of the summed mortality in the metalimninon and 88.5 to 94.2% of the summed mortality in the hypolimnion. Estimates of summed mortality values indicated that bacterial production was controlled completely or almost completely in the epilimnion (summed mortality, 66.6 to 128.5%) and the hypolimnion (summed mortality, 43.4 to 103.3%), whereas in the metalimnion viral lysis and HNF grazing were not sufficient to control bacterial production (summed mortality, 22.4 to 56.7%). The estimated contribution of organic matter released by viral lysis of cells into the pool of dissolved organic matter (DOM) was low; however, since cell lysis products are very likely labile compared to the bulk DOM, they might stimulate bacterial production. The high mortality of bacterioplankton due to viral lysis in anoxic water indicates that a significant portion of bacterial production in the metalimnion and hypolimnion is cycled in the bacterium-virus-DOM loop. This finding has major implications for the fate and cycling of organic nutrients in lakes.

Non-toluene-associated respiration in a Pseudomonas putida 54G biofilm grown on toluene in a flat-plate vapor-phase bioreactor

Non-toluene-associated respiration (NTAR) within a Pseudomonas putida 54G biofilm growing on toluene as sole external carbon source was evaluated using oxygen microelectrodes in a flat-plate vapor-phase biological reactor. Two fluorescent probes, 2,4-diamidino-2-phenylindole and 5-cyano-2,3-ditolyltetrazolium chloride, were used to evaluate the number of total and respiring cells respectively within the biofilm. Biofilm samples were also analyzed for viable and toluene-culturable cells by spread-plating on non-selective and selective media respectively. Fractions of viable stressed, respiring and non-respiring cells within the biofilm were evaluated. The NTAR rate was positively correlated with the fraction of viable stressed and non-respiring cells within the biofilm, which suggested the capability of some cells to grow at the expense of leakage and lysis products coming from injured and dead cells. This effect was more pronounced at higher toluene concentration. Results suggest that NTAR should be incorporated into mathematical models of biofilm reactors degrading volatile organic carbon compounds.

Comparison between Bone Marrow and G‐CSF‐Mobilized Peripheral Blood Allografts Undergoing Clinical Scale CD34 + Cell Selection

Allogeneic transplantation of selected CD34+ cells, rather than conventional transplantation of bone marrow (BM) harvest or peripheral blood (PB) leukapheresis products, has the advantage of reducing volume, facilitating storage and decreasing the amount of dimethylsulfoxide (DMSO) and cell lysis products, as well as reducing the number of T-lymphocytes responsible for graft-versus-host disease (GVHD). Using biotinavidin immunoaffinity columns (Ceprate SC system, CellPro; Bothell, WA), CD34+ cells were selected from each of 20 allografts (12 G-CSF-mobilized PB and 8 BM) collected from 14 HLA-identical normal healthy donors for transplantation. After the clinical-scale selection, the median concentration of CD34+ cells was 44.6% (range, 13% to 91%) in BM and 50.4% (range, 15% to 77%) in PB. Whereas 75% of the PB allografts had a CD34+ cell yield of more than 65%, only 37.5% of the BM allografts achieved such a yield, p < 0.01. The number of T-lymphocytes in the selected CD34+ cell allografts was reduced by two to three logs from a median of 4.2 x 10(9) to 7.8 x 10(5) CD3+ cells. The enrichment in CD34+ cells was 240-fold (range, 24- to 382-fold) in PB versus only 34-fold (range, 14- to 108-fold) in BM. Also, the enrichment in clonogenic cells was significantly more in PB (median value of 38.6-fold) than in BM (median value of 19.2-fold) and more in allografts from younger (< 50 years old) rather than older (> or = 50 years old) adult donors. A correlation was found between the percentage of CD34 or CD3+ cells before and after selection (r = 0.58 or r = 0.60, respectively, p < 0.05). Selective enrichment of the colony forming units-granulocyte-macrophage (CFU-GM) was found in all 20 allografts. The progenitor cell recovery after freezing and thawing was similar in BM and PB allografts, with a mean of about 60% for the CFU-GM and BFU-E. In the same six donors, the CD34+ cell yield was significantly more in the PB after mobilization (median 78.5%, range 50% to 90%) than in the BM before mobilization (median 41.5%, range 25% to 87%), p < 0.01. Ten patients with hematologic malignancies have been allotransplanted with 14 of the 20 selected CD34+ cells either combined BM + PB (n = 4) or single (n = 6) grafts. Seven patients did not develop acute GVHD, and only two patients developed > or = grade II GVHD, one of whom developed only grade II GVHD that resolved after brief treatment with corticosteriods. Only one patient showed chronic GVHD (skin and liver). The low incidence and severity of GVHD seen in the present study (only 30%) could be due to the two- to three-log reduction of T-lymphocytes in the selected CD34+ cell allotransplants. All 10 patients had stable hematological recovery, and seven had full donor hematopoiesis. In conclusion, G-CSF-mobilized PB leukapheresis products undergoing selection of CD34+ cells have a greater yield and enrichment of progenitor cells than BM harvests collected from HLA-identical normal healthy donors for allogeneic transplantation. The low incidence and severity of both acute and chronic GVHD (30%) seen in the present study are very encouraging.

Effects of viruses on nutrient turnover and growth efficiency of noninfected marine bacterioplankton

The effects of virus infection and lysis of a marine Vibrio sp. on C, N, and P turnover and the growth efficiency of noninfected bacterioplankton were studied in a series of dilution cultures. The cultures were enriched with various sources of organic matter and N and P. The growth of the Vibrio host and the growth of the natural bacterioplankton were measured by immunofluorescence and 4(prm1),6-diamidino-2-phenylindole staining methods, respectively. Lysis products resulting from infection of the Vibrio sp. caused an increase in metabolic activity and cell production by the noninfected bacterioplankton. In P-limited cultures, the addition of viruses increased the uptake of dissolved organic carbon by 72% and the potential alkaline phosphatase activity by 89% compared with control cultures without viruses. Our data suggest that input of available phosphorus through virus-induced Vibrio lysates occurred, which caused an increase in the bacterial nutrient uptake. The growth efficiency of noninfected bacteria was reduced in the presence of viruses compared with the control without viruses (growth efficiencies, 0.08 (plusmn) 0.03 and 0.24 (plusmn) 0.02, respectively). We suggest that the decrease in growth efficiency may be explained by an increase in bacterial energy demand associated with extracellular degradation of polymeric organic nitrogen and phosphorus in cell lysates.

Cell reabsorption in the Euphorbia dulcis endosperm

Euphorbia dulcis endosperm is the site of controlled long lasting endocellular lysis involving segregation and autophagy of portions of the cytoplasm within the endoplasmic reticulum membrane. The lysis products may constitute a food source for the benefit of the early developing embryo.

A more potent bystander cytocidal effect elicited by tumor cells expressing the herpes simplex virus-thymidine kinase gene than by fibroblast virus-producer cells in vitro.

Retrovirus-mediated herpes simplex virus-thymidine kinase (HSV-tk) gene therapy is a promising approach in the treatment of brain tumors. Previous in vitro and in vivo studies have demonstrated a bystander effect in which nonmodified tumor cells in proximity to HSV-tk-modified tumor cells are killed with the modified cells in the presence of ganciclovir. In the present study the authors assessed the contribution of infectious HSV-tk retrovirus made by producer cells to the bystander cytocidal effect in tissue culture using Walker 256 rat breast carcinosarcoma cells, which represent an established model for carcinomatous meningitis. The authors observed ganciclovir-dependent growth inhibition even when only one HSV-tk-positive Walker cell was mixed with 1000 HSV-tk-negative Walker cells and showed that the bystander cytocidal effect is not mediated by toxic cell lysis products. Walker cells engineered to produce HSV-tk retrovirus with titers ranging from 10(3) to 10(5) colony-forming units/ml exert no greater cytocidal effect than nonviral producer HSV-tk-positive Walker cells in vitro. Murine fibroblast-producer cells with viral titers ranging from 10(6) to 10(7) colony-forming units/ml exerted a stronger cytocidal effect than nonviral producer HSV-tk-positive murine fibroblasts. Despite the high viral titers of fibroblast producer cells, HSV-tk-modified Walker cells performed better than fibroblast producer cells in their cytotoxic effect on wild-type Walker tumor cells. Given that HSV-tk-modified tumor cells can become ganciclovir resistant, we tested gamma-irradiation as a means to overcome resistance. Lethal gamma-irradiation of the HSV-tk-positive Walker cells did not abolish their bystander effect on Walker HSV-tk-negative cells. One can infer from these results that HSV-tk-modified tumor cells, irradiated or not, may be a better alternative to murine fibroblast producer cells in the treatment of central nervous system neoplasia.

Decreased sludge production strategy for domestic wastewater treatment

With new EEC regulations, alternative treatment and disposal techniques of the excess sludge produced by Activated Sludge (AS) wastewater treatment plants have to be performed. In order to reduce the excess sludge produced, experiments have been carried out with a Membrane BioReactor (MBR) to study the maintenance and cryptic growth phenomena of Pseudomonas fluorescens culture taken as a model when grown on a limiting substrate complex medium similar to a synthetic urban wastewater. Experiments with various imposed wasting rates showed that viability and sludge production yield decreased when sludge age increased. Same variations were observed on the cell content ratio protein/polysaccharide by analysis of the cell lysis products released after discontinuous thermal treatment. Biomass growth on these cell lysis products was achieved to characterize cryptic growth and its impact on sludge production yield. Finally, a continuous sludge thermal treatment system was operating with MBR to amplify sludge breakage and consequently biomass growth on the lysis products. With the promising results obtained, this work gives a new outlook on the AS process and leads to the development of processes with control and reduction of sludge production.

Death and lysis during aerobic thermophilic sludge treatment: Characterization of recalcitrant products

The nature and fate of the organic fraction released during the aerobic thermophilic biodegradation of microbial cells were examined in a laboratory-scale treatment system. There was strong evidence for exoproteolytic activity during the early phase of the biodegradation process. The lysis products could be separated into two broad fractions, a low molecular weight C 2–C 5 carboxylic acid fraction and a pooled fraction of the remaining dissolved organic carbon (DOC). The former was produced as a result of fermentative metabolism of the aerobic thermophilic process culture growing under oxygen limited conditions. The production of these carboxylic acids occurred simultaneously with their utilization, as evidenced by the decrease in radioactivity in the DOC as a result of pulsing the culture with [ 14C]acetate. However, a fraction of this [ 14C]acetate pulse was metabolized into recalcitrant or slowly biodegradable DOC. The non-carboxylic acid fraction of the DOC in the lysis products was either slowly biodegradable or recalcitrant. The nature of the residual DOC in the supernatant after aerobic thermophilic treatment could not be further characterized but is likely to represent humic substances.

Algal bloom episodes and the formation of bituminite and micrinite in hydrocarbon source rocks: evidence from the Devonian and Mississippian, northern Williston Basin, Canada

Organic petrology (incident light microscopy) of Middle Devonian inter-reef laminates and Devonian-Mississippian epicontinental black shales, Williston Basin, Canada, indicates that algal bloom episodes and consequential bacterial activity played a significant role in the accumulation of amorphous, bituminite III-rich organic microfacies. Corpohuminite-like algal akinete cells produced by filamentous algae during algal bloom periods are persistent maceral inclusions within the potential hydrocarbon source rock intervals. These cells (% R o mean range 0.24-0.90) are regarded as positive indicators of stressful palaeoenvironmental conditions. Unicellular Tasmanites and Leiosphaeridia marine alginite and variably degraded alginite remnants (“ghosts”) within the amorphous kerogen may be products of cell lysis, photo-oxidation and microbial alteration; these processes are characteristic of algal bloom periods. Minute (ca. 1 μm) spheroidal and coccoidal bacteria-like macerals are dispersed throughout the bituminite III network, attesting to the importance of microbial activity within the water column and sediment during and after organic matter accumulation. Dispersed granules, laminations and replacement textures of micrinite-like macerals within bituminite III are interpreted as remnants of microbial alteration rather than a residual product of thermal maturation and hydrocarbon generation.

Quality of Blood Prepared for Autotransfusion in Primary Cemented Total Hip Replacement

We have measured levels of contaminants (products of cell lysis and other substances) in blood salvaged from patients undergoing primary cemented total hip replacement. Washing of this blood removed an average of 91.4% of the free haemoglobin mass, reduced white cell lysozyme concentrations by 86% and almost totally eliminated fats and particulate matter. Osmotic fragility was significantly improved by washing although not to control levels.

Clinical Toxicity of Cryopreserved Bone Marrow Graft Infusion

We prospectively evaluated infusion-related toxicities in 82 recipients of autologous bone marrow grafts. The grafts were cryopreserved in 10% dimethylsulfoxide and stored in liquid nitrogen. All grafts were concentrated and buffy-coat cells were collected. Forty-seven grafts were treated ex vivo with 4-hydroperoxycyclophosphamide (4-HC) at 100 micrograms/mL; 26 grafts were further processed using density-gradient separation and treated with 4-HC at 60 micrograms/mL. Nine buffy-coat concentrates were frozen without drug treatment. Before infusion, patients were medicated with mannitol, hydrocortisone, and diphenhydramine. Grafts were rapidly thawed and immediately infused without further manipulation. During the infusions, 33 (70%) recipients of treated buffy-coat, 5 (56%) recipients of untreated buffy-coat, and 6 (23%) recipients of density-gradient separated grafts experienced varying symptoms including nausea, abdominal cramping, and flushing. Forced vital capacities for 83% of the recipients of treated buffy-coat concentrates decreased after the graft infusion; six of these patients complained of dyspnea and one patient experienced an acute episode of respiratory decompensation. Decreased heart rates were observed in 98% of the recipients of treated buffy-coat cells with asymptomatic bradycardia occurring in 45%. Forty-five patients (96%) in this group experienced transient hypertension, with 18 (38%) requiring additional medications within 6 hours after the infusion for control of blood pressure. Similar cardiovascular changes were observed in the recipients of untreated buffy-coat concentrates. One recipient of an untreated buffy-coat concentrate had 2 degrees heart block after the graft infusion. Twenty-three (88%) recipients of density-gradient separated grafts had decreased heart rates and 21 (81%) had increased blood pressure. However, the degrees of change were less than those experienced by the recipients of treated buffy-coat cells (P less than .01). Forced vital capacities were not affected by the infusion of the density-gradient separated grafts. No renal failure or obvious hemolytic episodes occurred for any patient group. Minor to moderate toxicities were associated with cryopreserved graft infusions. Recipients of buffy-coat separated grafts, both treated and untreated, experienced more complications than the recipients of density-gradient separated grafts. These toxicities may relate to the volumes of cryoprotectant and cell lysis products infused, which were less for the more highly purified density-gradient separated grafts.