Potential use of whole cell lipase from a newly isolated Aspergillus nomius for methanolysis of palm oil to biodiesel

A highly active whole cell lipase (WCL) for efficient methanolysis of palm oil (PO) to biodiesel (BD) was prepared by isolation, cultivation and immobilization of lipase producing fungi. Fungi were screened from soil and the best isolate (PDA-6) identified as Aspergillus nomius exhibited maximum WCL methanolysis activity (1.4 g h⁻¹ g⁻¹) when inexpensive waste cooking oil was used as carbon source. The maximum BD yield with PDA-6 WCL reached 95.3% after 40 h at a lipase load 10% (w/w) of PO and methanol to PO molar ratio 5:1. The immobilization of PDA-6 cells within biomass suspended particle (BSP) made of polyurethane foam improved the repeated use of WCL and the remaining activity after 10 cycles was 88.2%. The PDA-6 WCL was more active in methanolysis of PO to BD than most WCLs previously reported. The newly isolated A. nomius is not only potential for producing WCL but also utilizing waste cooking oil.     

Morphological study of Cyclotella choctawhatcheeana Prasad (Stephanodiscaceae) from a saline Mexican lake

Background

Extracellular dissolved DNA has been demonstrated to be present in many terrestrial and aquatic environments, actively secreted, or released by decaying cells. Free DNA has the genetic potential to be acquired by living competent cells by horizontal gene transfer mediated by natural transformation. The aim of this work is to study the persistence of extracellular DNA and its biological transforming activity in extreme environments like the deep hypersaline anoxic lakes of the Mediterranean Sea. The brine lakes are separated from the upper seawater by a steep chemocline inhabited by stratified prokaryotic networks, where cells sinking through the depth profile encounter increasing salinity values and osmotic stress.

Results

Seven strains belonging to different taxonomic groups isolated from the seawater-brine interface of four hypersaline lakes were grown at medium salinity and then incubated in the brines. The osmotic stress induced the death of all the inoculated cells in variable time periods, between 2 hours and 144 days, depending on the type of brine rather than the taxonomic group of the strains, i.e. Bacillaceae or gamma-proteobacteria. The Discovery lake confirmed to be the most aggressive environment toward living cells. In all the brines and in deep seawater dissolved plasmid DNA was substantially preserved for a period of 32 days in axenic conditions. L'Atalante and Bannock brines induced a decrease of the supercoiled form up to 70 and 40% respectively; in the other brines only minor changes in plasmid conformation were observed. Plasmid DNA after incubation in the brines maintained the capacity to transform naturally competent cells of Acinetobacter baylii strain BD413.

Conclusion

Free dissolved DNA is likely to be released by the lysis of cells induced by osmotic stress in the deep hypersaline anoxic lakes. Naked DNA was demonstrated to be preserved and biologically active in these extreme environments, and hence could constitute a genetic reservoir of traits acquirable by horizontal gene transfer.     

Characterization of carp thrombocytes with specific monoclonal antibodies

Monoclonal antibodies (WCL6) specific for carp Cyprinus carpio thrombocytes were produced by immunizing mice with membrane lysates of IgM-negative peripheral blood leucocytes (PBL) and selected on the negative reaction with B cells. WCL6 was reactive with a membrane molecule of approximately 90 kDa and to a lesser extent with molecules of approximately 95 and 110 kDa. In general, between 30 and 40% of PBL were WCL6⁺ and appeared to be round to spindle-shaped cells. Immunohistochemical analysis of cryo-sections showed much higher numbers of WCL6⁺ cells in the spleen than in the pronephros, intestine and thymus. Flow cytometric analysis of cell suspensions isolated from these organs only revealed WCL6⁺ cells (4–10%) in the spleen. Electron microscopy of immunogold-stained WCL6⁺ PBL showed round but also some spindle-shaped cells with canalicular and granular structures, and a more irregular and electron-dense nucleus than found in lymphocytes. WCL6⁺ cells with electrondense pyknotic nuclei (without a clear nuclear envelope) were found also and their frequency increased with the length of the isolation and staining procedure used. In the spleen, several differentiation steps of WCL6⁺ cells were found and hence the spleen seems to be the thrombopoietic organ in carp. Thrombocytes from blood could be activated with collagen; the collagen-activated cells showed a higher side (90°) scatter by flow cytometric analysis and finally considerable cell death.     

Distribution and molecular evolution of rhamnose-binding lectins in Salmonidae: isolation and characterization of two lectins from white-spotted Charr (Salvelinus leucomaenis) eggs

L-Rhamnose-binding lectins were isolated from white-spotted charr (Salvelinus leucomaenis) eggs to understand the distribution and molecular evolution of the lectins in Salmonidae. Only two L-rhamnose-binding lectins, named WCL1 and WCL3, were isolated from white-spotted charr eggs, though three lectins, named STL1, STL2, and STL3, had been obtained from steelhead trout (Oncorhynchus mykiss) eggs. The cDNAs of WCL1 and WCL3 included 1,245 and 838 bp nucleotides with open reading frames of 933 and 651 nucleotides, respectively, and encoded for the complete amino acid sequences of mature proteins consisted of 288 (WCL1) and 195 (WCL3) residues, and signal sequences of 23 and 22 residues, respectively. WCLs were composed of three (for WCL1) or two (for WCL3) tandemly repeated homologous domains, which consisted of about 95 amino acid residues, and showed 91 and 93% sequence identities to STL1 and STL3, respectively. The mRNAs of WCL1 and WCL3 were detected exclusively in liver and ovary, respectively, however, neither a protein nor mRNA corresponding to STL2 could be identified in white-spotted charr. The phylogenetic tree of the sequences encoding carbohydrate recognition domains of 7 lectins from 4 species shows 5 functional clusters and their evolutional process. These results indicate that multiple L-rhamnose-binding isolectins have diverged by gene duplication and exon shuffling to play various biological roles in each species.     

East Tibetan Lakes Harbour Novel Clusters of Picocyanobacteria as Inferred from the 16S–23S rRNA Internal Transcribed Spacer Sequences

Planktonic picocyanobacteria abundance and diversity were investigated in nine lakes on the East Tibetan Plateau spanning a salinity gradient of 0.4–22.6 g l⁻¹. The investigation was conducted using epifluorescence microscopy (EFM) and terminal restriction fragment polymorphism analysis of 16S–23S rRNA internal transcribed spacer (ITS) PCR amplicons followed by sequence analyses of large ITS clone libraries of seven selected samples. EFM showed that picocyanobacteria comprised 7–19% of the total prokaryotic cells found in surface water. Most of the clones were classified into six clusters and grouped within the “picocyanobacterial clade”, which consists exclusively of freshwater Synechococcus. Four new phylogenetic clusters and one new subcluster of Synechococcus spp. were found, none of which are members of any known picocyanobacterial clusters. The new clusters and subcluster were the most abundant picocyanobacteria (about 96% of the sequences) in the samples collected. Sequence analyses indicated that members of the four new Synechococcus groups were only found in freshwater lakes (<1.0 g l⁻¹ of total dissolved solid), while members of the new subcluster were found in all the investigated Tibetan lakes, over a large salinity gradient of 0.4–22.6 g l ⁻¹. This suggests that there is ecologically significant microdiversity within the observed Synechococcus group as defined by ITS sequences. Collectively our study demonstrated abundant and potentially novel Synechococcus in East Tibetan lakes that are likely the result of evolutionary adaptations to regional conditions.     

Taxonomy of halophilic Archaea: current status and future challenges

Several groups of Archaea, all Euryarchaeota, develop in hypersaline environments (from >10 % salt up to saturation). The cultured diversity of halophilic Archaea includes the family Halobacteriaceae of aerobic or facultative anaerobic, generally red-pigmented species (47 genera and 165 species as of February 2014) and seven representatives of four genera of methanogens, most of which obtain energy from methylated amines under anaerobic conditions. Metagenomic studies have identified an additional deep lineage of Archaea in salt lakes and ponds with brines approaching NaCl saturation. Genomic information is now available for representatives of these ‘Nanohaloarchaea’, but no members of this lineage have yet been cultured. Multilocus sequence analysis is becoming increasingly popular in taxonomic studies of the Halobacteriaceae, and such studies have demonstrated that recombination of genetic traits occurs at an extremely high frequency at least in some genera. Metagenomic studies in an Antarctic lake showed that large identical regions of up to 35 kb in length can be shared by members of different genera living together in the same environment. Such observations have important implications not only for the taxonomy of the Halobacteriaceae, but also for species concepts and questions on taxonomy and classification for prokaryotic microorganisms in general.     

REMOVAL OF ORGANIC AND INORGANIC MATTER FROM ANTARCTIC LAKES BY AERIAL ESCAPE OF BLUEGREEN ALGAL MATS1

Field studies on three perennially ice-covered Antarctic oasis lakes with little or no outflow disclosed a unique biological phenomenon. Benthic algal mats dominated by the prokaryotic Phormidium frigidum Fritsch and several pennate diatoms growing in shallower, more brightly illuminated areas beneath 4 to 5.5 m of ice accumulate and entrap bubbles of photosynthetically produced oxygen. Clumps of this gas-filled mat tear loose from the gravelly substrate, lift off and float to the bottom of the lake ice. Some of these floating mat pieces become frozen into newly forming ice with the onset of winter. Through the combination of ablation of ice from the upper surface and the formation of new ice from below, algal mat pieces reach the upper lake surface in 5–10 years. Here, they are lyophilized by polar winds and dispersed in at least a partially viable state. The process of mat lift-off and escape is important in removing nutrients and salts from these lakes and helps to perpetuate their oligotrophic state. Neutron activation and X-ray dispersive analyses of elements in the algal mats along with other analyses, field observations, and calculations suggest that significant quantities of organic matter, select minerals and salts are lost from the lakes annually through this process whose magnitude has not been recognized previously.     

Sulfate-reducing bacteria in gypsum karst lakes of northern Lithuania

Microbiological studies were performed in three small gypsum karst lakes in northern Lithuania, most typical of the region. Samples were taken in different seasons of 2001. The conditions for microbial growth in the lakes are determined by elevated content of salts (from 0.5 to 2.0 g/l), dominated by SO(2-)4 and Ca2+ ions (up to 1.4 and 0.6 g/l, respectively). The elevated sulfate concentration is favorable for sulfate-reducing bacteria (SRBs). Summer and winter stratification gives rise to anaerobic water layers enriched in products of anaerobic degradation: H2S and CH4. The lakes under study contain abundant SRBs not only in bottom sediments (from 10(3) to 10(7) cells/dm3) but also in the water column (from 10(2) to 10(6) cells/ml). The characteristic spatial and temporal variations in the rate of sulfate reduction were noted. The highest rates of this process were recorded in summer: 0.95-2.60 mg S(2-)/dm3 per day in bottom sediments and up to 0.49 mg S(2-)/l per day in the water column. The maximum values (up to 11.36 mg S(2-)/dm3) were noted in areas where bottom sediments were enriched in plankton debris. Molecular analysis of conservative sequences of the gene for 16S RNA in sulfate-reducing microorganisms grown on lactate allowed them to be identified as Desulfovibrio desulfuricans.     

Prokaryotic diversity and community composition in the Salar de Uyuni,a large scale,chaotropic salt flat

Salar de Uyuni (SdU), with a geological history that reflects 50 000 years of climate change, is the largest hypersaline salt flat on Earth and is estimated to be the biggest lithium reservoir in the world. Its salinity reaches saturation levels for NaCl, a kosmotropic salt, and high concentrations of MgCL₂ and LiCl, both salts considered important chaotrophic stressors. In addition, extreme temperatures, anoxic conditions, high UV irradiance, high albedo and extremely low concentrations of phosphorous, make SdU a unique natural extreme environment in which to contrast hypotheses about limiting factors of life diversification. Geophysical studies of brines from different sampling stations show that water activity is rather constant along SdU. Geochemical measurements show significant differences in magnesium concentration, ranging from 0.2 to 2M. This work analyses the prokaryotic diversity and community structure at four SdU sampling stations, selected according to their location and ionic composition. Prokaryotic communities were composed of both Archaea (with members of the classes Halobacteria, Thermoplasmata and Nanohaloarchaea, from the Euryarchaeota and Nanohaloarcheota phyla respectively) and Bacteria (mainly belonging to Bacteroidetes and Proteobacteria phyla). The important differences in composition of microbial communities inversely correlate with Mg²⁺ concentration, suggesting that prokaryotic diversity at SdU is chaotropic dependent.     

Recent carbonate sedimentation and brine evolution in the saline lake basins of the Cariboo Plateau,British Columbia,Canada

There are more than 100 closed, saline lakes in the semiarid, intermontane plateaus of British Columbia. They range from shallow perennial lakes to ephemeral playas. Most are groundwater-fed and lie within glaciofluvial deposits and till. Some have permanent salts. Where underlain by basalts, sodium carbonate brines predominate. Magnesium sulphate brines occur where catchments lie within Paleozoic sedimentary rocks, metasediments and basic volcanics. A few sodium sulphate brines are also present.A reconnaissance study of the sediments and mineralogy of 21 lake basins has shown that carbonates, including extensive magnesite and hydromagnesite deposits, and several occurrences of protodolomite, are widely precipitated in lake basins of each brine type. Analyses of stream, spring, ground and lake waters from the Cariboo Plateau region demonstrate that carbonate precipitation probably constitutes the major chemical divide responsible for producing the two dominant types of brine.     

Effects of chaotropic salts on global proteome stability in halophilic archaea: Implications for life signatures on Mars

Halophilic archaea thriving in hypersaline environments, such as salt lakes, offer models for putative life in extraterrestrial brines such as those found on Mars. However, little is known about the effect of the chaotropic salts that could be found in such brines, such as MgCl₂, CaCl₂ and (per)chlorate salts, on complex biological samples like cell lysates which could be expected to be more representative of biomarkers left behind putative extraterrestrial life forms. We used intrinsic fluorescence to study the salt dependence of proteomes extracted from five halophilic strains: Haloarcula marismortui, Halobacterium salinarum, Haloferax mediterranei, Halorubrum sodomense and Haloferax volcanii. These strains were isolated from Earth environments with different salt compositions. Among the five strains that were analysed, H. mediterranei stood out as a results of its high dependency on NaCl for its proteome stabilization. Interestingly, the results showed contrasting denaturation responses of the proteomes to chaotropic salts. In particular, the proteomes of strains that are most dependent or tolerant on MgCl₂ for growth exhibited higher tolerance towards chaotropic salts that are abundant in terrestrial and Martian brines. These experiments bridge together global protein properties and environmental adaptation and help guide the search for protein-like biomarkers in extraterrestrial briny environments.     

Sulfate-Reducing Bacteria in Gypsum Karst Lakes of Northern Lithuania

Microbiological studies were performed in three small gypsum karst lakes in northern Lithuania, most typical of the region. Samples were taken in different seasons of 2001. The conditions for microbial growth in the lakes are determined by elevated content of salts (from 0.5 to 2.0 g/l), dominated by SO ₄ ^2? and Ca²⁺ ions (up to 1.4 and 0.6 g/l, respectively). The elevated sulfate concentration is favorable for sulfate-reducing bacteria (SRBs). Summer and winter stratification gives rise to anaerobic water layers enriched in products of anaerobic degradation: H₂S and CH₄. The lakes under study contain abundant SRBs not only in bottom sediments (from 10³ to 10⁷ cells/dm³) but also in the water column (from 10² to 10⁶ cells/ml). The characteristic spatial and temporal variations in the rate of sulfate reduction were noted. The highest rates of this process were recorded in summer: 0.95–2.60 mg S^2?/dm³ per day in bottom sediments and up to 0.49 mg S^2?/l per day in the water column. The maximum values (up to 11.36 mg S^2?/dm³) were noted in areas where bottom sediments were enriched in plankton debris. Molecular analysis of conservative sequences of the gene for 16S rRNA in sulfate-reducing microorganisms grown on lactate allowed them to be identified as Desulfovibrio desulfuricans.     

Ecological classification of lakes: Uncertainty and the influence of year-to-year variability

Regular monitoring of lakes is important to determine their ecological state and development and of key significance when deciding whether action should be taken to improve their quality, for instance by reducing the external loading of nutrients. Imprecise or inadequate knowledge of the ecological state increases the risk of misclassification and of wrong management decisions. Based on Danish lake data, we aimed to determine temporal variations, in particular natural year-to-year differences, and to describe the uncertainty in assessing the ecological state of lakes. We analysed environmental data from ca. 350 Danish lakes (1100 lake years), including three case studies, with long-term data series (up to 24 years), with no significant changes in external nutrient loading. We used summer means of selected water chemical variables, phytoplankton and submerged macrophytes as indicators of ecological state and found considerable variations in all indicators, which could not be ascribed alone to meteorological variation. In shallow lakes, chlorophyll a concentrations exhibited large year-to-year variations, especially at TP ranging between 0.05 and 0.15 mg L⁻¹ where the lakes may shift between a macrophyte- and a phytoplankton-dominated state. For example, chlorophyll a varied by a factor 5–10 between years and was particularly low when submerged macrophyte coverage exceeded 20% compared with lakes without macrophytes. Use of a multimetric index including four phytoplankton indicators reduced the coefficient of variation. Generally, the 95% confidence interval of ecological classification was approximately 50% lower when the assessment of ecological state was based on 4–5 years’ measurements than if based on only one year's measurements. Knowledge and awareness of the uncertainty of indicators used in ecological classification are highly relevant for lake managers and policy makers when defining efficient monitoring and restoration strategies.     

Comparative 16S rRNA analysis of lake bacterioplankton reveals globally distributed phylogenetic clusters including an abundant group of actinobacteria

In a search for cosmopolitan phylogenetic clusters of freshwater bacteria, we recovered a total of 190 full and partial 16S ribosomal DNA (rDNA) sequences from three different lakes (Lake Gossenk?llesee, Austria; Lake Fuchskuhle, Germany; and Lake Baikal, Russia). The phylogenetic comparison with the currently available rDNA data set showed that our sequences fall into 16 clusters, which otherwise include bacterial rDNA sequences of primarily freshwater and soil, but not marine, origin. Six of the clusters were affiliated with the alpha, four were affiliated with the beta, and one was affiliated with the gamma subclass of the Proteobacteria; four were affiliated with the Cytophaga-Flavobacterium-Bacteroides group; and one was affiliated with the class Actinobacteria (formerly known as the high-G+C gram-positive bacteria). The latter cluster (hgcI) is monophyletic and so far includes only sequences directly retrieved from aquatic environments. Fluorescence in situ hybridization (FISH) with probes specific for the hgcI cluster showed abundances of up to 1.7 x 10(5) cells ml(-1) in Lake Gossenk?llesee, with strong seasonal fluctuations, and high abundances in the two other lakes investigated. Cell size measurements revealed that Actinobacteria in Lake Gossenk?llesee can account for up to 63% of the bacterioplankton biomass. A combination of phylogenetic analysis and FISH was used to reveal 16 globally distributed sequence clusters and to confirm the broad distribution, abundance, and high biomass of members of the class Actinobacteria in freshwater ecosystems.     

Lake responses to reduced nutrient loading – an analysis of contemporary long-term data from 35 case studies

1. This synthesis examines 35 long‐term (5–35 years, mean: 16 years) lake re‐oligotrophication studies. It covers lakes ranging from shallow (mean depth <5 m and/or polymictic) to deep (mean depth up to 177 m), oligotrophic to hypertrophic (summer mean total phosphorus concentration from 7.5 to 3500 μg L^?1 before loading reduction), subtropical to temperate (latitude: 28–65°), and lowland to upland (altitude: 0–481 m). Shallow north‐temperate lakes were most abundant. 2. Reduction of external total phosphorus (TP) loading resulted in lower in‐lake TP concentration, lower chlorophyll a (chl a) concentration and higher Secchi depth in most lakes. Internal loading delayed the recovery, but in most lakes a new equilibrium for TP was reached after 10–15 years, which was only marginally influenced by the hydraulic retention time of the lakes. With decreasing TP concentration, the concentration of soluble reactive phosphorus (SRP) also declined substantially. 3. Decreases (if any) in total nitrogen (TN) loading were lower than for TP in most lakes. As a result, the TN : TP ratio in lake water increased in 80% of the lakes. In lakes where the TN loading was reduced, the annual mean in‐lake TN concentration responded rapidly. Concentrations largely followed predictions derived from an empirical model developed earlier for Danish lakes, which includes external TN loading, hydraulic retention time and mean depth as explanatory variables. 4. Phytoplankton clearly responded to reduced nutrient loading, mainly reflecting declining TP concentrations. Declines in phytoplankton biomass were accompanied by shifts in community structure. In deep lakes, chrysophytes and dinophytes assumed greater importance at the expense of cyanobacteria. Diatoms, cryptophytes and chrysophytes became more dominant in shallow lakes, while no significant change was seen for cyanobacteria. 5. The observed declines in phytoplankton biomass and chl a may have been further augmented by enhanced zooplankton grazing, as indicated by increases in the zooplankton : phytoplankton biomass ratio and declines in the chl a : TP ratio at a summer mean TP concentration of <100–150 μg L^?1. This effect was strongest in shallow lakes. This implies potentially higher rates of zooplankton grazing and may be ascribed to the observed large changes in fish community structure and biomass with decreasing TP contribution. In 82% of the lakes for which data on fish are available, fish biomass declined with TP. The percentage of piscivores increased in 80% of those lakes and often a shift occurred towards dominance by fish species characteristic of less eutrophic waters. 6. Data on macrophytes were available only for a small subsample of lakes. In several of those lakes, abundance, coverage, plant volume inhabited or depth distribution of submerged macrophytes increased during oligotrophication, but in others no changes were observed despite greater water clarity. 7. Recovery of lakes after nutrient loading reduction may be confounded by concomitant environmental changes such as global warming. However, effects of global change are likely to run counter to reductions in nutrient loading rather than reinforcing re‐oligotrophication.     

Sedimentology and geochemistry of saline lakes of the Great Plains

Southern Saskatchewan and portions of adjacent Alberta, North Dakota and Montana are occupied by hundreds of saline and hypersaline lakes ranging in size from small prairie potholes (less than 1 km²) to relatively large bodies of water (greater than 300 km²). From a sedimentological perspective, distinction must be made between two basic types of saline lakes: playas and perennial lakes. Calcium, sodium and magnesium sulfates, carbonates and bicarbonates form as chemical precipitates in lakes with more concentrated brines. In addition, experimental data suggests mixed layer smectites may form authigenically in some lakes. Clastic sediments in the salt lakes consist mainly of silt and clay-sized quartz, feldspars, carbonates and clay minerals. The dominant physical and chemical processes which are responsible for and act upon the sediment vary widely, mainly in response to basin morphology and brine chemistry. Evaporative concentration and significant groundwater contributions affect all the saline lakes. However, other processes are different in the two basic types of basins. The playa lakes are influenced by: evaporitic pumping and the formation of efflorescent crusts and intrasedimentary crystals, cyclic wetting and drying, precipitation of highly soluble salt layers, and influx of clastic debris by sheetflood and wind. In contrast, in the permanent lakes, precipitation of sparingly soluble salts occurs due to the interaction of biological activity, seasonal temperature fluctuations and brine mixing. In addition, many of the permanent lakes undergo freeze-out precipitation of very soluble salts under a winter ice cover. Detrital sediments are distributed within the basins by normal lacustrine processes, including shoreline deposition and erosion, turbidity flow and pelagic fallout.     

Picoplankton in Six New Zealand Lakes: Abundance in Relation to Season and Trophic State

The abundance of picoplankton (0.2-2 μm) was measured seasonally in the surface waters of six New Zealand lakes that represent a range of trophic states. The lakes were: Wakatipu, Te Anau, Manapouri, Hayes, Mahinerangi and Ross Creek Reservoir. Among the lakes, picoplankton abundance was associated positively with temperature; picoplankton were most abundant in summer and autumn when they attained densities of 108,000-270,000 cells/ml in the oligotrophic lakes. In these lakes, prokaryotic picoplankton was generally an order of magnitude more abundant than eukaryotic picoplankton. Consistent with the hypothesis that picoplankton are more important in oligotrophic than eutrophic ecosystems, there was a weak negative correlation between the density of prokaryotic picoplankton in the lakes and the level of chlorophyll a. The presence of large numbers of chroococcoid cyanobacteria in the guts of Ceriodaphnia dubia and Bosmina meridionalis implies that prokaryotic picoplankton are collected, but not digested, by these species.     

BL-01, an Fc-bearing,tetravalent CD20 × CD5 bispecific antibody,redirects multiple immune cells to kill tumors in vitro and in vivo

Background aimsThe authors describe here a novel therapeutic strategy combining a bispecific antibody (bsAb) with cytokine-induced killer (CIK) cells.MethodsThe authors have designed, produced and purified a novel tetravalent IgG1-like CD20 × CD5 bsAb called BL-01. The bsAb is composed of a fused heavy chain and two free light chains that pair correctly to the heavy chain sequences thanks to complementary mutations in the monoclonal antibody 2 CH1/CL sequences.ResultsThe authors show that BL-01 can bind specifically to CD20 and CD5 with an affinity of 4–6 nM, demonstrating correct pairing of two light chains to the fused heavy chain. The CD20 × CD5 BL-01 bsAb has a functional human IgG1 Fc and can induce up to 65% complement-dependent cytotoxicity of a CD20⁺ lymphoma cell line in the presence of human complement, similar to anti-CD20 rituximab. The bsAb also induces significant natural killer cell activation and antibody-dependent cytotoxicity of up to 25% as well as up to 65% phagocytosis by human macrophages in the presence of CD20⁺ tumor cells. The BL-01 bsAb binds to CD20 and CD5 simultaneously and can redirect CIK cells in vitro to kill CD20⁺ targets, increasing the cytotoxicity of CIK cells by about 3-fold. The authors finally show that the CD20 × CD5 BL-01 bsAb synergizes with CIK cells in vivo in controlling tumor growth and prolonging survival of nonobese diabetic/severe combined immunodeficiency mice inoculated with a patient-derived, aggressive diffuse large B-cell lymphoma xenograft.ConclusionsThe authors suggest that the efficacy of bsAb in vivo is due to the combined activation of innate immunity by Fc and redirection of CIK cells to kill the tumor target.     

Microbial decomposition of organic matter in the bottom sediments of small lakes of the urban landscape (Lithuania)

Bacterial abundance and the rates of sulfate reduction (SR) and total organic matter decomposition (D_total) were studied in the bottom sediments of nine lakes in the vicinity of Vilnius (Lithuania) during the ice-free seasons of 2006–2009. During the spring mixing of the water, aerobic processes of organic matter decomposition prevailed in the bottom sediments of most lakes, while anaerobic processes predominated (up to 80–90% D_total) in summer and early autumn. SR rates in the bottom sediments made up 0.16–2.6 and 0.09–2.0 mg S^2?/(dm³ day) for the medium-depth and shallow lakes, respectively. The highest numbers of sulfate-reducing bacteria (up to 10⁶ cells/cm³) and SR rates were observed in summer. SR rate in mediumdepth lakes increased with development of anaerobic conditions at the bottom and elevated sulfate concentrations (up to 96.0 mg/dm³). In shallow lakes, where O₂ concentration at the bottom was at least 6.7 mg/L, SR rates increased with temperature and inflow of fresh organic matter, especially during cyanobacterial blooms. The average SR rates in the bottom sediments of the lakes of urbanized areas were 4 times higher than in the shallow lakes of protected areas. Accumulation of organic matter and its intensive decomposition during summer may enhance the processes of secondary eutrophication of these small and shallow lakes.