Growth factors involved in lymphocyte differentiation

We report here that an interleukin-3-dependent precursor B-cell line, LyD9, differentiated in vitro into mature B cells, producing immunoglobulin (Ig)M and IgG by co-culture with bone marrow stroma cells. Induced LyD9 cells underwent heterogenous immunoglobulin gene rearrangement and synthesized mRNAs encoding immunoglobulin mu (mu), gamma (gamma) and kappa (kappa) chains. LyD9 was also shown to differentiate into myeloid cells. We have established an interleukin-4-dependent derivative clone K-4 that is an intermediate between myeloid-lymphoid cells and the LyD9 clone. This differentiation required direct contact between LyD9 and stromal cells.     

Active autotrophic ammonia-oxidizing bacteria in biofilm enrichments from simulated creek ecosystems at two ammonium concentrations respond to temperature manipulation

The first step of nitrification, the oxidation of ammonia to nitrite, is important for reducing eutrophication in freshwater environments when coupled with anammox (anaerobic ammonium oxidation) or denitrification. We analyzed active formerly biofilm-associated aerobic ammonia-oxidizing communities originating from Ammerbach (AS) and Leutra South (LS) stream water (683 ± 550 [mean ± standard deviation] and 16 ± 7 μM NH(4)(+), respectively) that were developed in a flow-channel experiment and incubated under three temperature regimens. By stable-isotope probing using (13)CO(2), we found that members of the Bacteria and not Archaea were the functionally dominant autotrophic ammonia oxidizers at all temperatures under relatively high ammonium loads. The copy numbers of bacterial amoA genes in (13)C-labeled DNA were lower at 30°C than at 13°C in both stream enrichment cultures. However, the community composition of the ammonia-oxidizing bacteria (AOB) in the (13)C-labeled DNA responded differently to temperature manipulation at two ammonium concentrations. In LS enrichments incubated at the in situ temperature (13°C), Nitrosomonas oligotropha-like sequences were retrieved with sequences from Nitrosospira AmoA cluster 4, while the proportion of Nitrosospira sequences increased at higher temperatures. In AS enrichments incubated at 13°C and 20°C, AmoA cluster 4 sequences were dominant; Nitrosomonas nitrosa-like sequences dominated at 30°C. Biofilm-associated AOB communities were affected differentially by temperature at two relatively high ammonium concentrations, implicating them in a potential role in governing contaminated freshwater AOB distributions.     

Growth and stability of ethene-utilizing bacteria on compost at very low substrate concentrations

Ethene-utilising micro-organisms on compost may be applied in a packed bed to scrub the plant hormone ethene from air. Ethene at the concentrations tested (50–200 vpm) supported growth of micro-organisms present in compost and also of ethene-grown Mycobacterium E3 cells immobilised on compost.     

Growth of ammonia-oxidizing archaea and bacteria in cattle manure compost under various temperatures and ammonia concentrations

A recent study showed that ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) coexist in the process of cattle manure composting. To investigate their physiological characteristics, liquid cultures seeded with fermenting cattle manure compost were incubated at various temperatures (37°C, 46°C, or 60°C) and ammonium concentrations (0.5, 1, 4, or 10?mM NH (4) (+) -N). The growth rates of the AOB and AOA were monitored using real-time polymerase chain reaction analysis targeting the bacterial and archaeal ammonia monooxygenase subunit A genes. AOB grew at 37°C and 4 or 10?mM NH (4) (+) -N, whereas AOA grew at 46°C and 10?mM NH (4) (+) -N. Incubation with allylthiourea indicated that the AOB and AOA grew by oxidizing ammonia. Denaturing gradient gel electrophoresis and subsequent sequencing analyses revealed that a bacterium related to Nitrosomonas halophila and an archaeon related to Candidatus Nitrososphaera gargensis were the predominant AOB and AOA, respectively, in the seed compost and in cultures after incubation. This is the first report to demonstrate that the predominant AOA in cattle manure compost can grow and can probably oxidize ammonia under moderately thermophilic conditions.     

Nitrate-based niche differentiation by distinct sulfate-reducing bacteria involved in the anaerobic oxidation of methane

Diverse associations between methanotrophic archaea (ANME) and sulfate-reducing bacterial groups (SRB) often co-occur in marine methane seeps; however, the ecophysiology of these different symbiotic associations has not been examined. Here, we applied a combination of molecular, geochemical and Fluorescence in situ hybridization (FISH) coupled to nanoscale secondary ion mass spectrometry (FISH-NanoSIMS) analyses of in situ seep sediments and methane-amended sediment incubations from diverse locations (Eel River Basin, Hydrate Ridge and Costa Rican Margin seeps) to investigate the distribution and physiology of a newly identified subgroup of the Desulfobulbaceae (seepDBB) found in consortia with ANME-2c archaea, and compared these with the more commonly observed associations between the same ANME partner and the Desulfobacteraceae (DSS). FISH analyses revealed aggregates of seepDBB cells in association with ANME-2 from both environmental samples and laboratory incubations that are distinct in their structure relative to co-occurring ANME/DSS consortia. ANME/seepDBB aggregates were most abundant in shallow sediment depths below sulfide-oxidizing microbial mats. Depth profiles of ANME/seepDBB aggregate abundance revealed a positive correlation with elevated porewater nitrate relative to ANME/DSS aggregates in all seep sites examined. This relationship with nitrate was supported by sediment microcosm experiments, in which the abundance of ANME/seepDBB was greater in nitrate-amended incubations relative to the unamended control. FISH-NanoSIMS additionally revealed significantly higher ¹⁵N-nitrate incorporation levels in individual aggregates of ANME/seepDBB relative to ANME/DSS aggregates from the same incubation. These combined results suggest that nitrate is a geochemical effector of ANME/seepDBB aggregate distribution, and provides a unique niche for these consortia through their utilization of a greater range of nitrogen substrates than the ANME/DSS.     

Dynamics of specific ammonia-oxidizing bacterial populations and nitrification in response to controlled shifts of ammonium concentrations in wastewater

Ammonia-oxidizing bacteria (AOB) are essential for the nitrification process in wastewater treatment. To retain these slow-growing bacteria in wastewater treatment plants (WWTPs), they are often grown as biofilms, e.g., on nitrifying trickling filters (NTFs) or on carriers in moving bed biofilm reactors (MBBRs). On NTFs, a decreasing ammonium gradient is formed because of the AOB activity, resulting in low ammonium concentrations at the bottom and reduced biomass with depth. To optimize the NTF process, different ammonium feed strategies may be designed. This, however, requires knowledge about AOB population dynamics. Using fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy, we followed biomass changes during 6 months, of three AOB populations on biofilm carriers. These were immersed in aerated MBBR tanks in a pilot plant receiving full-scale wastewater. Tanks were arranged in series, forming a wastewater ammonium gradient mimicking an NTF ammonium gradient. The biomass of one of the dominating Nitrosomonas oligotropha-like populations increased after an ammonium upshift, reaching levels comparable to the high ammonium control in 28 days, whereas a Nitrosomonas europaea-like population increased relatively slowly. The MBBR results, together with competition studies in NTF systems fed with wastewater under controlled ammonium regimes, suggest a differentiation between the two N. oligotropha populations, which may be important for WWTP nitrification.     

Epiphyton as a niche for ammonia-oxidizing bacteria: detailed comparison with benthic and pelagic compartments in shallow freshwater lakes

Next to the benthic and pelagic compartments, the epiphyton of submerged macrophytes may offer an additional niche for ammonia-oxidizing bacteria in shallow freshwater lakes. In this study, we explored the potential activities and community compositions of ammonia-oxidizing bacteria of the epiphytic, benthic, and pelagic compartments of seven shallow freshwater lakes which differed in their trophic status, distribution of submerged macrophytes, and restoration history. PCR-denaturing gradient gel electrophoresis analyses demonstrated that the epiphytic compartment was inhabited by species belonging to cluster 3 of the Nitrosospira lineage and to the Nitrosomonas oligotropha lineage. Both the ammonia-oxidizing bacterial community compositions and the potential activities differed significantly between compartments. Interestingly, both the ammonia-oxidizing bacterial community composition and potential activity were influenced by the restoration status of the different lakes investigated.     

Effects of ammonium and nitrite on communities and populations of ammonia-oxidizing bacteria in laboratory-scale continuous-flow reactors

This study investigated the effects of ammonium and nitrite on ammonia-oxidizing bacteria (AOB) from an activated sludge process in laboratory-scale continuous-flow reactors. AOB communities were analyzed using specific PCR followed by denaturing gel gradient electrophoresis, cloning and sequencing of the 16S rRNA gene, and AOB populations were quantified using real-time PCR. To study the effect of ammonium, activated sludge from a sewage treatment system was enriched in four reactors receiving inorganic medium containing four different ammonium concentrations (2, 5, 10 and 30 mM NH(4) (+)-N). One of several sequence types of the Nitrosomonas oligotropha cluster predominated in the reactors with lower ammonium loads (2, 5 and 10 mM NH(4) (+)-N), whereas Nitrosomonas europaea was the dominant AOB in the reactor with the highest ammonium load (30 mM NH(4) (+)-N). The effect of nitrite was studied by enriching the enriched culture possessing both N. oligotropha and N. europaea in four reactors receiving 10-mM-ammonium inorganic medium containing four different nitrite concentrations (0, 2, 12 and 22 mM NO(2) (-)-N). Nitrosomonas oligotropha comprised the majority of AOB populations in the reactors without nitrite accumulation (0 and 2 mM NO(2) (-)-N), whereas N. europaea was in the majority in the 12- and 22-mM NO(2) (-)-N reactors, in which nitrite concentrations were 2.1-5.7 mM (30-80 mg N L(-1)).     

Growth of bacteria degrading naphthalene and salicylate at low temperatures

A total of 58 bacterial strains degrading naphthalene and salicylate were isolated from soil samples polluted with oil products, collected in different regions of Russia during winter and summer. The isolates were assessed for their ability to grow at low temperatures (4, 8, and 15 degrees C); bacteria growing at 4 degrees C in the presence of naphthalene or salicylate accounted for 65% and 53%, respectively, of the strains isolated. The strains differed in the temperature dependence of their growth rates. It was demonstrated that the type of expression of Nah+ phenotype at low temperatures depended on the combination of the host bacterium and the plasmid.     

Quantification of ammonia-oxidizing bacteria and factors controlling nitrification in salt marsh sediments

To elucidate the geomicrobiological factors controlling nitrification in salt marsh sediments, a comprehensive approach involving sediment geochemistry, process rate measurements, and quantification of the genetic potential for nitrification was applied to three contrasting salt marsh habitats: areas colonized by the tall (TS) or short (SS) form of Spartina alterniflora and unvegetated creek banks (CBs). Nitrification and denitrification potential rates were strongly correlated with one another and with macrofaunal burrow abundance, indicating that coupled nitrification-denitrification was enhanced by macrofaunal burrowing activity. Ammonia monooxygenase (amoA) gene copy numbers were used to estimate the ammonia-oxidizing bacterial population size (5.6 x 10(4) to 1.3 x 10(6) g of wet sediment(-1)), which correlated with nitrification potentials and was 1 order of magnitude higher for TS and CB than for SS. TS and CB sediments also had higher Fe(III) content, higher Fe(III)-to-total reduced sulfur ratios, higher Fe(III) reduction rates, and lower dissolved sulfides than SS sediments. Iron(III) content and reduction rates were positively correlated with nitrification and denitrification potential and amoA gene copy number. Laboratory slurry incubations supported field data, confirming that increased amounts of Fe(III) relieved sulfide inhibition of nitrification. We propose that macrofaunal burrowing and high concentrations of Fe(III) stimulate nitrifying bacterial populations, and thus may increase nitrogen removal through coupled nitrification-denitrification in salt marsh sediments.     

Competition and coexistence of aerobic ammonium- and nitrite-oxidizing bacteria at low oxygen concentrations

In natural and man-made ecosystems nitrifying bacteria experience frequent exposure to oxygen-limited conditions and thus have to compete for oxygen. In several reactor systems (retentostat, chemostat and sequencing batch reactors) it was possible to establish co-cultures of aerobic ammonium- and nitrite-oxidizing bacteria at very low oxygen concentrations (2–8 μM) provided that ammonium was the limiting N compound. When ammonia was in excess of oxygen, the nitrite-oxidizing bacteria were washed out of the reactors, and ammonium was converted to mainly nitrite, nitric oxide and nitrous oxide by Nitrosomonas-related bacteria. The situation could be rapidly reversed by adjusting the oxygen to ammonium ratio in the reactor. In batch and continuous tests, no inhibitory effect of ammonium, nitric oxide or nitrous oxide on nitrite-oxidizing bacteria could be detected in our studies. The recently developed oxygen microsensors may be helpful to determine the kinetic parameters of the nitrifying bacteria, which are needed to make predictive kinetic models of their competition.     

Cell surface properties as factors involved in Proteus vulgaris adhesion to stainless steel under starvation conditions

The purpose of these investigations was to evaluate the influence of limited nutrient availability in the culture medium on Proteus vulgaris biofilm formation on surfaces of stainless steel. The relationship between the P. vulgaris adhesion to the abiotic surfaces, the cellular ATP levels, cell surface hydrophobicity and changes in the profiles of extracellular proteins and lipopolysaccharides was examined. In all experimental variants the starvation conditions induced the bacterial cells to adhere to the surfaces of stainless steel. Higher ATP content and lower cell surface hydrophobicity of P. vulgaris cells was observed upon nutrient-limited conditions. Under starvation conditions a reduction in the levels of extracellular low molecular weight proteins was noticed. High molecular weight proteins formed the conditioning layer on stainless steel plates, making the bacteria adhesion process more favorable. The production of low molecular weight carbohydrates promoted more advanced stages of P. vulgaris biofilm formation process on the surfaces of stainless steel upon starvation.     

Degradation of 2,3,4,6-tetrachlorophenol at low temperature and low dioxygen concentrations by phylogenetically different groundwater and bioreactor bacteria

Effects of low temperature and low oxygen partial pressure on theoccurrence and activity of 2,3,4,6-tetrachlorophenol degrading bacteria in a boreal chlorophenol contaminated groundwater and a full-scale fluidized-bed bioreactor were studied using four polychlorophenol degrading bacterial isolates of different phylogenetic backgrounds. These included an -proteobacterial Sphingomonas sp. strain MT1 isolated from the full-scale bioreactor and three isolates from the contaminated groundwater whichwere identified as -proteobacterial Herbaspirillum sp. K1,a Gram-positive bacterium with high G + C content Nocardioides sp. K44 and an -proteobacterialSphingomonas sp. K74. The Sphingomonasstrains K74 and MT1 and Nocardioides sp. K44 degraded2,4,6-trichlorophenol and 2,3,4,6-tetrachlorophenol as the solecarbon and energy sources. Close to stoichiometric inorganic chloride release with the 2,3,4,6-tetrachlorophenol removal andthe absence of methylation products indicated mineralization. Tetrachlorophenol degradation by the Herbaspirillum sp. K1 was enhanced by yeast extract, malate, glutamate, pyruvate, peptone and casitone. At 8 °C, Sphingomonas sp. K74 had the highest specific degradation rate(_max = 4.9 × 10^-12 mg h^-1 cell^-1) for 2,3,4,6-tetrachlorophenol. The Nocardioides strain K44 had the highest affinity (K_s = 0.46 mg l^-1) for tetrachlorophenol. K1 and MT1 grew microaerophilically in semisolid glucose medium. Furthermore, the growth of MT1 was inhibited in liquidglucose medium at high oxygen partial pressure indicating sensitivity to accumulating toxic oxygen species. On the other hand, trichlorophenol degradation was not affected by oxygen concentration (2–21%). The isolates K44, K74 and MT1, with optimum growth temperaturesbetween 23 and 25 °C, degraded tetrachlorophenol faster at 8 °C than at room temperature indicating distinctly different temperature optima for chlorophenol degradation and growthon complex media. These results show efficient polychlorophenol degradation by the isolates at the boreal groundwater conditions, i.e., at low temperature and low oxygen concentrations. Differences in chlorophenol degradation and sensitivities to chlorophenols and oxygen among the isolates indicate that the phylogenetically different chlorophenol degraders have found different niches in the contaminated groundwater and thus potential for contaminantdegradation under a variety of saturated subsurface conditions.     

Growth and uptake kinetics of a facultatively oligotrophic bacterium at low nutrient concentrations

In oligotrophic waters, not only community structure but also physiological properties of heterotrophic bacteria are influenced by the concentration of organic matter.The relationship between growth rate of two facultatively oligotrophic strains ofAeromonas sp. No. 6 andFlavobacterium sp. M1 was studied in comparison with that of two eutrophic strains ofEscherichia coli 7020 andFlavobacterium sp. M2. These strains had two or three different substrate constants (Ks values) depending on substrate concentrations: Ks values for the two former were remarkably lower than those for the two latter. For instance, Ks value forAeromonas sp. No. 6 was about 8.9M when substrate concentration was greater than 53M and about 1.1M when substrate concentration was less man 53M. InE. coli the Ks value was about 260M at greater than 5600M and about 47M at less than 5600M substrate concentration.Uptake kinetics ofAeromonas sp. grown in a medium containing 2.7 mM glutamate (H-cell) and 0.11M glutamate (L-cell) have been determined for the intact cells. H-cell had two distinct values of Km for glutamate assimilation and respiration, and L-cell had three distinct values of Km for glutamate assimilation and respiration: In H-cell Km of assimilation was 2.8×10^–7 M and 1.5×10^–4 M, and Km of respiration was 2.3×10^–7 M and 1.7×10^–4 M; in L-cell Km of assimilation was 7.4×10^–8 M, 8.3×10^–6 M, and 1.3×10^–4 M, and Km of respiration was 2.5×10^–7, 8.9×10^–6M, and 1.7×10^–4 M. More than 60% of glutamate taken up by the H- and L-cells was respired when the substrate concentration was less than 10^–6 M, although at greater than 10^–6 M, 50% and 30% of glutamate was respired by H-cells and L-cells, respectively. These results suggest that the facultatively oligotrophic bacteria grow with high efficiency in environments with extremely low nutrient concentration, such as oligotrophic waters of lakes and ocean, as compared with in their growth in conditions of high nutrient concentraton, such as nutrient broth.     

Systems utilizing oxygenated leghemoglobin and myoglobin as sources of free dissolved O2 at low concentrations for experiments with bacteria

Oxygenated soybean leghemoglobin and sperm whale myoglobin have been used as sources of O₂ for respiring bacteria in experiments with no gas phase. These O₂-carrying hemoproteins provide dispersed free O₂ at concentrations defined by the kinetic constants of their oxygenation and deoxygenation, and their optical absorption spectra indicate the average concentration of free O₂. They can thus be used to study relationships between concentrations of free O₂, rates of respiration, and other metabolic processes in bacteria suspended in solutions of them. Three types of apparatus are described and examples of studies of respiration and nitrogen fixation by Rhizobium spp. are given.