The P2CS database (http://www.p2cs.org/) is a comprehensive resource for the analysis of Prokaryotic Two-Component Systems (TCSs). TCSs are comprised of a receptor histidine kinase (HK) and a partner response regulator (RR) and control important prokaryotic behaviors. The latest incarnation of P2CS includes 164 651 TCS proteins, from 2758 sequenced prokaryotic genomes. Several important new features have been added to P2CS since it was last described. Users can search P2CS via BLAST, adding hits to their cart, and homologous proteins can be aligned using MUSCLE and viewed using Jalview within P2CS. P2CS also provides phylogenetic trees based on the conserved signaling domains of the RRs and HKs from entire genomes. HK and RR trees are annotated with gene organization and domain architecture, providing insights into the evolutionary origin of the contemporary gene set. The majority of TCSs are encoded by adjacent HK and RR genes, however, 'orphan' unpaired TCS genes are also abundant and identifying their partner proteins is challenging. P2CS now provides paired HK and RR trees with proteins from the same genetic locus indicated. This allows the appraisal of evolutionary relationships across entire TCSs and in some cases the identification of candidate partners for orphan TCS proteins.
TiO2-based nanocomposite (NC) are widely used as invisible UV protectant in cosmetics. These nanomaterials (NMs) end in the environment as altered materials. We have investigated the properties of T-Lite SF, a TiO2-NC used as sunscreen, after weathering in water and under light. We have examined the formation of ROS and their consequences on cell physiology of Escherichia coli. Our results show that aged-T-Lite SF produced singlet oxygen under low intensity long wave UV and formed hydroxyl radicals at high intensity. Despite the production of these ROS, T-Lite SF had neither effect on the viability of E. coli nor on mutant impaired in oxidative stress, did not induce mutagenesis and did not impair the integrity of membrane lipids, thus seemed safe to bacteria. However, when pre-exposed to T-Lite SF under low intensity UV, cells turned out to be more sensitive to cadmium, a priority pollutant widely disseminated in soil and surface waters. This effect was not a Trojan horse: sensitization of cells was dependent on the formation of singlet oxygen. These results provide a basis for caution, especially on NMs that have no straight environmental toxicity. It is crucial to anticipate indirect and combined effects of environmental pollutants and NMs.
Microbial anodes were formed under polarization at -0.3 V/SCE on graphite plates in effluents from a pulp and paper mill. The bioanodes formed with the addition of acetate led to the highest current densities (up to 6A/m(2)) but were then unable to oxidize the raw effluent efficiently (0.5A/m(2)). In contrast, the bioanodes formed without acetate addition were fully able to oxidize the organic matter contained in the effluent, giving up to 4.5A/m(2) in continuous mode. Bacterial communities showed less bacterial diversity for the acetate-fed bioanodes compared to those formed in raw effluents. Deltaproteobacteria were the most abundant taxonomic group, with a high diversity for bioanodes formed without acetate addition but with almost 100% Desulfuromonas for the acetate-fed bioanodes. The addition of acetate to form the microbial anodes induced microbial selection, which was detrimental to the treatment of the raw effluent.
Physical-chemists, (micro)biologists, and ecologists need to conduct meaningful experiments to study the environmental risk of engineered nanomaterials with access to relevant mechanistic data across several spatial and temporal scales. Indoor aquatic mesocosms (60L) that can be tailored to virtually mimic any ecosystem appear as a particularly well-suited device. Here, this concept is illustrated by a pilot study aimed at assessing the distribution of a CeO?-based nanomaterial within our system at low concentration (1.5 mg/L). Physico-chemical as well as microbiological parameters took two weeks to equilibrate. These parameters were found to be reproducible across the 9-mesocosm setup over a 45-day period of time. Recovery mass balances of 115 ± 18% and 60 ± 30% of the Ce were obtained for the pulse dosing and the chronic dosing, respectively. This demonstrated the relevance of our experimental approach that allows for adequately monitoring the fate and impact of a given nanomaterial.
Garden compost leachate was used to form microbial bioanodes under polarization at -0.4, -0.2 and +0.1 V/SCE. Current densities were 6.3 and 8.9 A m(-2) on average at -0.4 and +0.1 V/SCE respectively, with acetate 10 mM. The catalytic cyclic voltammetry (CV) showed similar electrochemical characteristics for all bioanodes and indicated that the lower currents recorded at -0.4V/SCE were due to the slower interfacial electron transfer rate at this potential, consistently with conventional electrochemical kinetics. RNA- and DNA-based DGGE evidenced that the three dominant bacterial groups Geobacter, Anaerophaga and Pelobacter were identical for all bioanodes and did not depend on the polarization potential. Only non-turnover CVs showed differences in the redox equipment of the biofilms, the highest potential promoting multiple electron transfer pathways. This first description of a potential-independent electroactive microbial community opens up promising prospects for the design of stable bioanodes for microbial fuel cells.
Heavy metals such as cadmium (Cd(2+)) affect microbial metabolic processes. Consequently, bacteria adapt by adjusting their cellular machinery. We have investigated the dose-dependent growth effects of Cd(2+) on Rhizobium alamii, an exopolysaccharide (EPS)-producing bacterium that forms a biofilm on plant roots. Adsorption isotherms show that the EPS of R. alamii binds cadmium in competition with calcium. A metabonomics approach based on ion cyclotron resonance Fourier transform mass spectrometry has showed that cadmium alters mainly the bacterial metabolism in pathways implying sugars, purine, phosphate, calcium signalling and cell respiration. We determined the influence of EPS on the bacterium response to cadmium, using a mutant of R. alamii impaired in EPS production (MS?GT). Cadmium dose-dependent effects on the bacterial growth were not significantly different between the R. alamii wild type (wt) and MS?GT strains. Although cadmium did not modify the quantity of EPS isolated from R. alamii, it triggered the formation of biofilm vs planktonic cells, both by R. alamii wt and by MS?GT. Thus, it appears that cadmium toxicity could be managed by switching to a biofilm way of life, rather than producing EPS. We conclude that modulations of the bacterial metabolism and switching to biofilms prevails in the adaptation of R. alamii to cadmium. These results are original with regard to the conventional role attributed to EPS in a biofilm matrix, and the bacterial response to cadmium.
This study investigated the influence of uranium on the indigenous bacterial community structure in natural soils with high uranium content. Radioactive soil samples exhibiting 0.26% - 25.5% U in mass were analyzed and compared with nearby control soils containing trace uranium. EXAFS and XRD analyses of soils revealed the presence of U(VI) and uranium-phosphate mineral phases, identified as sabugalite and meta-autunite. A comparative analysis of bacterial community fingerprints using denaturing gradient gel electrophoresis (DGGE) revealed the presence of a complex population in both control and uranium-rich samples. However, bacterial communities inhabiting uraniferous soils exhibited specific fingerprints that were remarkably stable over time, in contrast to populations from nearby control samples. Representatives of Acidobacteria, Proteobacteria, and seven others phyla were detected in DGGE bands specific to uraniferous samples. In particular, sequences related to iron-reducing bacteria such as Geobacter and Geothrix were identified concomitantly with iron-oxidizing species such as Gallionella and Sideroxydans. All together, our results demonstrate that uranium exerts a permanent high pressure on soil bacterial communities and suggest the existence of a uranium redox cycle mediated by bacteria in the soil.
Ramlibacter tataouinensis TTB310(T) (strain TTB310), a betaproteobacterium isolated from a semi-arid region of South Tunisia (Tataouine), is characterized by the presence of both spherical and rod-shaped cells in pure culture. Cell division of strain TTB310 occurs by the binary fission of spherical "cyst-like" cells ("cyst-cyst" division). The rod-shaped cells formed at the periphery of a colony (consisting mainly of cysts) are highly motile and colonize a new environment, where they form a new colony by reversion to cyst-like cells. This unique cell cycle of strain TTB310, with desiccation tolerant cyst-like cells capable of division and desiccation sensitive motile rods capable of dissemination, appears to be a novel adaptation for life in a hot and dry desert environment. In order to gain insights into strain TTB310s underlying genetic repertoire and possible mechanisms responsible for its unusual lifestyle, the genome of strain TTB310 was completely sequenced and subsequently annotated. The complete genome consists of a single circular chromosome of 4,070,194 bp with an average G+C content of 70.0%, the highest among the Betaproteobacteria sequenced to date, with total of 3,899 predicted coding sequences covering 92% of the genome. We found that strain TTB310 has developed a highly complex network of two-component systems, which may utilize responses to light and perhaps a rudimentary circadian hourglass to anticipate water availability at the dew time in the middle/end of the desert winter nights and thus direct the growth window to cyclic water availability times. Other interesting features of the strain TTB310 genome that appear to be important for desiccation tolerance, including intermediary metabolism compounds such as trehalose or polyhydroxyalkanoate, and signal transduction pathways, are presented and discussed.
To shed light on the genetic equipment of the beneficial plant-associated bacterium Pseudomonas brassicacearum, we sequenced the whole genome of the strain NFM421. Its genome consists of one chromosome equipped with a repertoire of factors beneficial for plant growth. In addition, a complete type III secretion system and two complete type VI secretion systems were identified. We report here the first genome sequence of this species.
A specificity of Brassicaceous plants is the production of sulphur secondary metabolites called glucosinolates that can be hydrolysed into glucose and biocidal products. Among them, isothiocyanates are toxic to a wide range of microorganisms and particularly soil-borne pathogens. The aim of this study was to investigate the role of glucosinolates and their breakdown products as a factor of selection on rhizosphere microbial community associated with living Brassicaceae. We used a DNA-stable isotope probing approach to focus on the active microbial populations involved in root exudates degradation in rhizosphere. A transgenic Arabidopsis thaliana line producing an exogenous glucosinolate and the associated wild-type plant associated were grown under an enriched (13)CO(2) atmosphere in natural soil. DNA from the rhizospheric soil was separated by density gradient centrifugation. Bacterial (Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria and Acidobacteria), Archaea and fungal community structures were analysed by DGGE fingerprints of amplified 16S and 18S rRNA gene sequences. Specific populations were characterized by sequencing DGGE fragments. Roots of the transgenic plant line presented an altered profile of glucosinolates and other minor additional modifications. These modifications significantly influenced microbial community on roots and active populations in the rhizosphere. Alphaproteobacteria, particularly Rhizobiaceae, and fungal communities were mainly impacted by these Brassicaceous metabolites, in both structure and composition. Our results showed that even a minor modification in plant root could have important repercussions for soil microbial communities.
Electrochemically active biofilms were constructed on graphite anodes under constant polarization at -0.1V vs saturated calomel reference (SCE) with 10 mM acetate as substrate. The reactors were inoculated with three different microbial samples that were drawn from exactly the same place in a French Atlantic coastal port (i) by scraping the biofilm that had formed naturally on the surface of a floating bridge, (ii) by taking marine sediments just under the floating bridge, and (iii) by taking nearby beach sand. Current densities of 2.0 A/m2 were reached using the biofilm sample as inoculum while only 0.4 A/m2 and 0.8 A/m2 were obtained using the underlying sediments and the beach sand, respectively. The structure of bacterial communities forming biofilms was characterized by denaturing gradient gel electrophoresis (DGGE) analysis, and revealed differences between samples with the increase in relative intensities of some bands and the appearance of others. Bacteria close related to Bacteroidetes, Halomonas, and Marinobacterium were retrieved only from the efficient EA-biofilms formed from natural biofilms, whereas, bacteria close related to Mesoflavibacter were predominant on biofilm formed from sediments. The marine biofilm was selected as the inoculum to further optimize the microbial anode. Epifluorescence microscopy and SEM confirmed that maintaining the electrode under constant polarization promoted rapid settlement of the electrode surface by a bacterial monolayer film. The microbial anode was progressively adapted to the consumption of acetate by three serial additions of substrate, thus improving the Coulombic efficiency of acetate consumption from 31 to 89%. The possible oxidation of sulfide played only a very small part in the current production and the biofilm was not able to oxidize hydrogen. Graphite proved to be more efficient than dimensionally stable anode (DSA) or stainless steel butthis result might be due to differences in the surface roughness rather than the intrinsic features of the materials. Finally, a maximal current density of 7.9 A/m2 was reached with 10 mM acetate after only 8 days of biofilm formation at -0.1 V/SCE. These results are among the best performance values reported in the literature. Using natural biofilms as inoculum should, consequently, be a new, very promising wayto rapidly build more efficient microbial electrodes than those produced when the inoculum is drawn from bulk environments.
Five isolates obtained from a marine electroactive biofilm grown on a stainless steel cathode were investigated by using a polyphasic taxonomic approach. Analyses of whole-cell fatty acid methyl esters and 16S rRNA gene sequences showed that the isolates belonged to the Roseobacter lineage of the class Alphaproteobacteria. Both phenotypic and genotypic analyses demonstrated that the five new isolates constituted a single species that did not represent a recognized member of the Roseobacter lineage. Therefore the five isolates represent a novel genus and species, for which the name Nautella italica gen. nov., sp. nov. is proposed. The type strain is LMG 24365(T) (=CCUG 55857(T)). The DNA G+C content of the type strain is 61 mol%.
To better understand adaptation to harsh conditions encountered in hot arid deserts, we report the first complete genome sequence and proteome analysis of a bacterium, Deinococcus deserti VCD115, isolated from Sahara surface sand. Its genome consists of a 2.8-Mb chromosome and three large plasmids of 324 kb, 314 kb, and 396 kb. Accurate primary genome annotation of its 3,455 genes was guided by extensive proteome shotgun analysis. From the large corpus of MS/MS spectra recorded, 1,348 proteins were uncovered and semiquantified by spectral counting. Among the highly detected proteins are several orphans and Deinococcus-specific proteins of unknown function. The alliance of proteomics and genomics high-throughput techniques allowed identification of 15 unpredicted genes and, surprisingly, reversal of incorrectly predicted orientation of 11 genes. Reversal of orientation of two Deinococcus-specific radiation-induced genes, ddrC and ddrH, and identification in D. deserti of supplementary genes involved in manganese import extend our knowledge of the radiotolerance toolbox of Deinococcaceae. Additional genes involved in nutrient import and in DNA repair (i.e., two extra recA, three translesion DNA polymerases, a photolyase) were also identified and found to be expressed under standard growth conditions, and, for these DNA repair genes, after exposure of the cells to UV. The supplementary nutrient import and DNA repair genes are likely important for survival and adaptation of D. deserti to its nutrient-poor, dry, and UV-exposed extreme environment.
A group of exopolysaccharide-producing bacteria was isolated from the root environment of Arabidopsis thaliana. The genetic diversity revealed by REP-PCR fingerprinting indicated that the isolates correspond to different strains. 16S rRNA gene sequence analysis showed that the isolates are closely related to the strains Rhizobium sp. YAS34 and USDA 1920, respectively isolated from sunflower roots and Medicago ruthenica nodules. These bacteria belong to the Rhizobium lineage of the Alphaproteobacteria, and the closest known species was Rhizobium sullae. DNA-DNA hybridization experiments and biochemical analysis demonstrated that the nine strains isolated from A. thaliana and Rhizobium strains YAS34 and USDA 1920 constitute a novel species within the genus Rhizobium, for which the name Rhizobium alamii sp. nov. is proposed. The type strain is GBV016(T) (=CFBP 7146(T) =LMG 24466(T)).
Stainless steel electrodes were cathodically polarized at -200 mV versus an Ag/AgCl reference electrode in natural seawater in order to produce current. The current increased and stabilized at 0.5 A/m(2) in less than 10 days. The cultivable fraction of the microbial biofilm population formed on the surface of the current harvesting cathodes was examined by culture dependent techniques. Three hundred fifty six isolates were obtained. They were primarily characterized by whole cell fatty acid methyl ester analysis followed by 16S rRNA gene sequencing. The results showed that the isolates represented different phylogenetic groups including members of the Alpha- and Gammaproteobacteria, the phylum Firmicutes, the family Flavobacteriacae and the phylum Actinobacteria. Denaturing gradient gel electrophoresis demonstrated that the microbial population of the biofilm formed on the cathode and of the surrounding seawater comprised the same dominant members. This study demonstrated that the cultivable microbial fraction of a marine electroactive biofilm is phylogenetically highly diverse.
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Hydrogen production by microorganisms is often described as a promising sustainable and clean energy source, but still faces several obstacles, which prevent practical application. Among them, oxygen sensitivity of hydrogenases represents one of the major limitations hampering the biotechnological implementation of photobiological production processes. Here, we describe a hierarchical biodiversity-based approach, including a chemochromic screening of hydrogenase activity of hundreds of bacterial strains collected from several ecosystems, followed by mass spectrometry measurements of hydrogenase activity of a selection of the H(2)-oxidizing bacterial strains identified during the screen. In all, 131 of 1266 strains, isolated from cereal rhizospheres and basins containing irradiating waste, were scored as H(2)-oxidizing bacteria, including Pseudomonas sp., Serratia sp., Stenotrophomonas sp., Enterobacter sp., Rahnella sp., Burkholderia sp., and Ralstonia sp. isolates. Four free-living N(2)-fixing bacteria harbored a high and oxygen-tolerant hydrogenase activity, which was not fully inhibited within entire cells up to 150-250 ?mol/L O(2) concentration or within soluble protein extracts up to 25-30 ?mol/L. The only hydrogenase-related genes that we could reveal in these strains were of the hyc type (subunits of formate hydrogenlyase complex). The four free-living N(2)-fixing bacteria were closely related to Enterobacter radicincitans based on the sequences of four genes (16S rRNA, rpoB, hsp60, and hycE genes). These results should bring interesting prospects for microbial biohydrogen production and might have ecophysiological significance for bacterial adaptation to the oxic-anoxic interfaces in the rhizosphere.
Microbial anodes were formed under polarisation at -0.2V/SCE on smooth graphite plate electrodes with paper mill effluents. Primary, secondary and tertiary biofilms were formed by a successive scratching and re-inoculation procedure. The secondary and tertiary biofilms formed while decreasing the polarisation potential allowed the anodes to provide current density of 6A/m(2) at -0.4V/SCE. In contrast, applying -0.4V/SCE initially to form the primary biofilms did not lead to the production of current. Consequently, the scratching/re-inoculation procedure combined with progressive lowering of the applied potential revealed an efficient new procedure that gave efficient microbial anodes able to work at low potential. The observed progressive pH drift to alkaline values above 9 explained the open circuit potentials as low as -0.6 V/SCE. The remarkable performance of the electrode at alkaline pH was attributed to the presence of Desulfuromonas acetexigens as the single dominant species in the tertiary microbial anodes.
Electrochemically active (EA) biofilms were formed on metallic dimensionally stable anode-type electrode (DSA), embedded in garden compost and polarized at +0.50 V/SCE. Analysis of 16S rRNA gene libraries revealed that biofilms were heavily enriched in Deltaproteobacteria in comparison to control biofilms formed on non-polarized electrodes, which were preferentially composed of Gammaproteobacteria and Firmicutes. Among Deltaproteobacteria, sequences affiliated with Pelobacter and Geobacter genera were identified. A bacterial consortium was cultivated, in which 25 isolates were identified as Geobacter bremensis. Pure cultures of 4 different G. bremensis isolates gave higher current densities (1400 mA/m(2) on DSA, 2490 mA/m(2) on graphite) than the original multi-species biofilms (in average 300 mA/m(2) on DSA) and the G. bremensis DSM type strain (100-300 A/m(2) on DSA; 2485 mA/m(2) on graphite). FISH analysis confirmed that G. bremensis represented a minor fraction in the original EA biofilm, in which species related to Pelobacter genus were predominant. The Pelobacter type strain did not show EA capacity, which can explain the lower performance of the multi-species biofilms. These results stressed the great interest of extracting and culturing pure EA strains from wild EA biofilms to improve the current density provided by microbial anodes.
The rhizosphere is an active compartment where plant and microorganisms establish a molecular dialogue. In this study, we analysed the impact of Arabidopsis thaliana on bacterial community structure and the expression of certain beneficial genes using DNA- and mRNA-SIP in the rhizosphere of plantlets grown under (13)CO(2) for 13, 21 and 27 days. DNA- and rRNA-SIP revealed changes in bacterial communities inhabiting the rhizosphere soil that were probably related to modification of root exudates, while root-colonizing populations were maintained over time suggesting their metabolic versatility and adaptation. The impact of the plant via root exudates on the expression of the noncoding RNAs rsmZ, acdS gene encoding 1-aminocyclopropane-1-carboxylate deaminase and nosZ gene encoding nitrous oxide reductase, in the root-adhering soil and on the roots of A. thaliana was determined using mRNA-SIP. Results showed that these genes were present and expressed by bacteria inhabiting roots and by those that derive nutrients from the breakdown of organic matter in soils or from root exudates. The expression of rsmZ under natural conditions indicates the importance of noncoding RNAs in bacterial adaptation to their ecological niches.
The plant-beneficial bacterium Pseudomonas brassicacearum forms phenotypic variants in vitro as well as in planta during root colonization under natural conditions. Transcriptome analysis of typical phenotypic variants using microarrays containing coding as well as noncoding DNA fragments showed differential expression of several genes relevant to secondary metabolism and of the small RNA (sRNA) genes rsmX, rsmY, and rsmZ. Naturally occurring mutations in the gacS-gacA system accounted for phenotypic switching, which was characterized by downregulation of antifungal secondary metabolites (2,4-diacetylphloroglucinol and cyanide), indoleacetate, exoenzymes (lipase and protease), and three different N-acyl-homoserine lactone molecules. Moreover, in addition to abrogating these biocontrol traits, gacS and gacA mutations resulted in reduced expression of the type VI secretion machinery, alginate biosynthesis, and biofilm formation. In a gacA mutant, the expression of rsmX was completely abolished, unlike that of rsmY and rsmZ. Overexpression of any of the three sRNAs in the gacA mutant overruled the pleiotropic changes and restored the wild-type phenotypes, suggesting functional redundancy of these sRNAs. In conclusion, our data show that phenotypic switching in P. brassicacearum results from mutations in the gacS-gacA system.
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