Anaerobic benzene degradation under denitrifying conditions: Peptococcaceae as dominant benzene degraders and evidence for a syntrophic process

An anaerobic microbial community was enriched in a chemostat that was operated for more than 8 years with benzene and nitrate as electron acceptor. The coexistence of multiple species in the chemostat and the presence of a biofilm, led to the hypothesis that benzene-degrading species coexist in a syntrophic interaction, and that benzene can be degraded in syntrophy by consortia with various electron acceptors in the same culture. The benzene-degrading microorganisms were identified by DNA-stable isotope probing with [U-(13) C]-labelled benzene, and the effect of different electron donors and acceptors on benzene degradation was investigated. The degradation rate constant of benzene with nitrate (0.7 day(-1) ) was higher than reported previously. In the absence of nitrate, the microbial community was able to use sulfate, chlorate or ferric iron as electron acceptor. Bacteria belonging to the Peptococcaceae were identified as dominant benzene consumers, but also those related to Rhodocyclaceae and Burkholderiaceae were found to be associated with the anaerobic benzene degradation process. The benzene degradation activity in the chemostat was associated with microbial growth in biofilms. This, together with the inhibiting effect of hydrogen and the ability to degrade benzene with different electron acceptors, suggests that benzene was degraded via a syntrophic process.     

Genome sequences of Alicycliphilus denitrificans strains BC and K601T

Alicycliphilus denitrificans strain BC and A. denitrificans strain K601(T) degrade cyclic hydrocarbons. These strains have been isolated from a mixture of wastewater treatment plant material and benzene-polluted soil and from a wastewater treatment plant, respectively, suggesting their role in bioremediation of soil and water. Although the strains are phylogenetically closely related, there are some clear physiological differences. The hydrocarbon cyclohexanol, for example, can be degraded by strain K601(T) but not by strain BC. Furthermore, both strains can use nitrate and oxygen as an electron acceptor, but only strain BC can use chlorate as electron acceptor. To better understand the nitrate and chlorate reduction mechanisms coupled to the oxidation of cyclic compounds, the genomes of A. denitrificans strains BC and K601(T) were sequenced. Here, we report the complete genome sequences of A. denitrificans strains BC and K601(T).     

Halorespiring bacteria-molecular characterization and detection

Recently, a rapidly increasing number of bacteria has been isolated that is able to couple the reductive dehalogenation of various halogenated aromatic and aliphatic compounds like chlorophenols and tetrachloroethene to energy conservation by electron-transport-coupled phosphorylation. The potential of these halorespiring bacteria for innovative clean-up strategies of polluted anoxic environments has greatly stimulated efforts to unravel the molecular basis of the novel respiratory chains they possess. The thorough characterization of halorespiratory key components at the physiological, biochemical and molecular genetic level has revealed both structural and functional similarity of chloroaryl- and chloroalkyl-respiratory chains from different phylogenetically distinct microorganisms. The reductive dehalogenases from halorespiring bacteria were found to comprise a novel class of corrinoid-containing Fe/S-proteins. Sensitive molecular methods for monitoring both presence and fate of halorespiring bacteria have been developed, which will be instrumental for the design and maintenance of optimised in situ bioremediation processes.     

Direct Profiling of Environmental Microbial Populations by Thermal Dissociation Analysis of Native rRNAs Hybridized to Oligonucleotide Microarrays

Oligonucleotide microarrays were used to profile directly extracted rRNA from environmental microbial populations without PCR amplification. In our initial inspection of two distinct estuarine study sites, the hybridization patterns were reproducible and varied between estuarine sediments of differing salinities. The determination of a thermal dissociation curve (i.e., melting profile) for each probe-target duplex provided information on hybridization specificity, which is essential for confirming adequate discrimination between target and nontarget sequences.     

Conjugative transfer, stability and expression of a plasmid encoding a cry1Ac gene in Bacillus cereus group strains

The plasmid pHT73 containing cry1Ac and tagged with an erythromycin resistance gene was transferred from Bacillus thuringiensis subspecies kurstaki KT0 to several Bacillus cereus group strains by conjugation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and phase contrast microscopy showed that the transconjugants containing plasmid pHT73 could express Cry1Ac toxin and produce bipyramidal crystalline inclusion bodies during sporulation. The study demonstrated that pHT73 could be transferred to B. thuringiensis subsp. kurstaki, several B. cereus strains and Bacillus mycoides. Under non-selective conditions, the stability of the pHT73 plasmid in the transconjugants was found to be 58.2-100% after 100 generations and 4-96% after 200 generations. The variations are mainly caused by the choice of receptor strain.     

Virgin forests in Romania and Bulgaria: results of two national inventory projects and their implications for protection

Despite extensive forest destruction in the Middle Ages and later intensive commercial forest management, remnants of virgin forests remained spared in some Central, Eastern and South-Eastern European countries. These virgin forests are the last examples of original forests in this part of Europe. That is why their protection becomes an important issue of current European forestry and nature protection policy. But the knowledge about the location and the area of virgin forests in these countries is incomplete up till now. This article has the prime goal to present a conceptual framework what virgin forests might be (“A conceptual framework for defining of virgin forests” section). Based on this framework, a working methodology has been tested in Bulgaria and Romania (“Results of the two national projects in Romania and in Bulgaria” section and further). For this reason two projects have been carried out by the Royal Dutch Society of Nature Conservation (KNNV) in close co-operation with the Forestry Institutes in Romania and in Bulgaria. The results of these projects are described in general terms and further analysis in the future is necessary to describe specific features like forest structure and spatial heterogeneity of these forests. Based on the results of the inventory, principles of sustainable protection and management of the mapped virgin forests were defined and described in the research reports. The usefulness of the inventory became evident already during the EU pre-accession period of both countries while preparing the NATURA 2000 network. The remaining virgin forests of temperate Europe are an inexhaustible source of ecological information about biodiversity, structure, natural processes and overall functioning of undisturbed forest ecosystems. Their research will reveal information which can be used for ecological restoration of man-made forests which are degraded through intensive forestry practices over the last centuries. The last virgin forests of temperate Europe represent an irreplaceable part of the natural capital of Europe and are worth to be protected by law. Their last remnants in South-Eastern and Eastern Europe are endangered by commercial activities. A full inventory of remaining virgin forests in all countries of temperate Europe is a matter of highest urgency. A representative selection of virgin forest sites should be declared by UNESCO as World Heritage Sites.     

Changes in dormancy of Sisymbrium officinaleseeds do not depend on changes in respiratory activity

Effects of dark incubation at different temperatures were studied on dormancy and respiratory activity of seeds of Sisymbrium officinale (L.) Scop. Because germination of this species absolutely depends on the simultaneous action of light and nitrate, changes in dormancy could be studied in darkness without the interference of early germination events. Upon the start of incubation rates of O₂ uptake and CO₂ release rose. This was followed by a gradual decrease until stable levels of O₂ uptake and CO₂ release were achieved. Seeds kept for prolonged periods at 24^°C, showed neither a change in germination capacity nor in rates of O₂ uptake and CO₂ release. Respiratory quotients were 0.55–0.7. The initial rise in O₂ uptake correlated with the rate of water uptake and with breaking of primary dormancy. However, the subsequent decline in O₂ uptake was not generally linked to induction of secondary dormancy. An increased O₂ uptake was not required during breaking of secondary dormancy. It is concluded that changes in dormancy are not generally related to changes in respiratory activity. However, germination strongly depends on respiration. The increase in O₂ uptake started well before radicle protrusion. A far red irradiation only reversed this increase when it was given before germination escaped from its red light antagonising action. The contribution of different respiratory pathways was followed during prolonged incubation at 24^°C in darkness. KCN at 1.5 mM was needed to inhibit the cytochrome pathway (CP) and benzohydroxamic acid (BHAM) at 30 mM to inhibit the alternative pathway (AP). These concentrations did not exert any side effects. Electron flow was predominantly via the CP, maximally 10% was via the AP. Flow through the CP declined during the first 6 days and residual respiration remained constant. Therefore, the contribution of residual respiration became relatively more important with prolonged incubation. KCN at concentrations that almost completely inhibited flow through the CP, did not dramatically reduce germination. BHAM already inhibited germination at concentrations that do not inhibit oxygen uptake.