39 research outputs found
Die Rolle der im Regenwurmdarm assoziierten Mikroorganismen auf das Schicksal des im Boden auftretenden Prions
The effect of earthworm gut (species Lumbricus terrestris, Aporrectodea caliginosa, and Eisenia fetida) on transiting bacterial community was studied using 16S rRNA-based techniques. CFB bacteria (classes Flavobacteria and Sphingobacteria) were considered as facultative gut-associated. Newly detected family Lumbricoplasmataceae within the class Mollicutes (Firmicutes) was proposed to be obligate earthworm-associated bacteria. Firmicutes and Gammaproteobacteria strongly reduced their diversity and relative number upon the passage being gut-sensitive bacterial groups, although the genus Pseudomonas (Gammaproteobacteria) were gut-resistant members of community. Other bacterial groups have varied number; unclassified Spingomonadaceae (Alphaproteobacteria) and Alcaligenes faecalis (Betaproteobacteria) represented the gut-resistant part of their populations. No earthworm host-specific effect was observed. Up to 20% of bacterial species isolated from the soil and earthworm sources were able to digest recPrP in pure culture; Gammaproteobacteria, Actinobacteria, and Bacilli were the taxa with the biggest potential to deplete the recPrP. Most of studied fungal isolates degraded recPrP. The prion was demonstrated to be depleted in vitro in aqueous extracts of the soil and the cast within 2-6 days. Non-specific proteolytic activity strongly increased from soil substratum to the cast through the trypsin- and chymotrypsin-like proteases released by the earthworm. However, the passage through the gut did not promote any enhanced recPrP digestion. Thus, under applied conditions the microbial-earthworm gut systems do not produce proteases de novo, which notably affect the prion proteolysis.Die Auswirkung der Regenwurmdarm (Arten Lumbricus terrestris, Aporrectodea caliginosa, und Eisenia fetida) assoziierten und durchgehenden Bakterien en wurde mit Hilfe der 16S rRNA-Bestimmung untersucht. CFB Bakterien (Klassen Flavobacteria und Sphingobacteria) wurden als fakultativ regenwurmdarmassoziiert betrachtet. Die neulich entdeckte Familie Lumbricoplasmataceae mit der Klasse Mollicutes (Firmicutes) wurde als regenwurmdarmassoziierte Bakterien vorgeschlagen. Bei Firmicutes und Gammaproteobacteria wurde eine starke Reduktion ihrer DiversitĂ€t, sowie der Zahl der Bakterien innerhalb der Darmpassage beobachtet, was auf eine darmempfindliche Bakteriengruppe hinweist, obwohl die Gattung Pseudomonas (Gammaproteobacteria) eine DarmwiederstandsfĂ€higkeit zeigte. Die Bakterienzahl der anderen Bakteriengruppen hat sich geĂ€ndert (variiert); nicht klassifizierte Spingomonadaceae (Alphaproteobacteria) und Alcaligenes faecalis (Betaproteobacteria) stellten einen darm-wiederstandsfĂ€higen Teil ihrer Population dar. Es wurde keine wirtspezifische Rolle des Regenwurms beobachtet. Es wurde keine spezifische Wirkung beobachtet bei RegenwĂŒrmern aus unterschiedlichen Umgebungen. Bis 20 % der aus dem Boden und den RegenwĂŒrmern isolierten Bakterienspezies waren dazu fĂ€hig, recPrP aus einer reinen Kultur abzubauen; Gammaproteobacteria, Actinobacteria, und Bacilli wiesen einen gröĂeres Potential zum Abbau von recPrP auf. Die meisten getesteten pilzartigen Isolate waren auch zum recPrP-Abbau fahig. Die Degradation des Prions wurde in vitro in den wĂ€ssrigen Extrakten aus dem Boden und den Wurmexkrementen (2-6 Tagen) demonstriert (gezeigt). Es wurde eine stark erhöhte unspezifische proteolytische AktivitĂ€t durch den in Regenwurmdarm (freigesetzten) detektierten trypsin- und chymotrypsin-artigen Enzymen beobachtet. Jedoch wurde kein erhöhte recPrP-Abbau innerhalb der Darmpassage detektiert. Zusammenfassend erzeugt das Mikrooraganismen-Regenwurmdarm System unter angewandten Bedingungen, keine de novo Protease, die zum Prion-Abbau beitragen
Metabolic and evolutionary patterns in the extremely acidophilic archaeon Ferroplasma acidiphilum YT
Ferroplasmaceae represent ubiquitous iron-oxidising extreme acidophiles with a number of unique physiological traits. In a genome-based study of Ferroplasma acidiphilum YT, the only species of the genus Ferroplasma with a validly published name, we assessed its central metabolism and genome stability during a long-term cultivation experiment. Consistently with physiology, the genome analysis points to F. acidiphilum YT having an obligate peptidolytic oligotrophic lifestyle alongside with anaplerotic carbon assimilation. This narrow trophic specialisation abridges the sugar uptake, although all genes for glycolysis and gluconeogenesis, including bifunctional unidirectional fructose 1,6-bisphosphate aldolase/phosphatase, have been identified. Pyruvate and 2-oxoglutarate dehydrogenases are substituted by âancientâ CoA-dependent pyruvate and alpha-ketoglutarate ferredoxin oxidoreductases. In the lab culture, after ~550 generations, the strain exhibited the mutation rate of â„1.3âĂâ10â8 single nucleotide substitutions per site per generation, which is among the highest values recorded for unicellular organisms. All but one base substitutions were G:C to A:T, their distribution between coding and non-coding regions and synonymous-to-non-synonymous mutation ratios suggest the neutral drift being a prevalent mode in genome evolution in the lab culture. Mutations in nature seem to occur with lower frequencies, as suggested by a remarkable genomic conservation in F. acidiphilum YT variants from geographically distant populations
âARMANâ archaea depend on association with euryarchaeal host in culture and in situ
AbstractIntriguing, yet uncultured âARMANâ-like archaea are metabolically dependent on other members of the microbial community. It remains uncertain though which hosts they rely upon, and, because of the lack of complete genomes, to what extent. Here, we report the co-culturing of ARMAN-2-related organism, Mia14, with Cuniculiplasma divulgatum PM4 during the isolation of this strain from acidic streamer in Parys Mountain (Isle of Anglesey, UK). Mia14 is highly enriched in the binary culture (ca. 10% genomic reads) and its ungapped 0.95âMbp genome points at severe voids in central metabolic pathways, indicating dependence on the host, C. divulgatum PM4. Analysis of C. divulgatum isolates from different sites and shotgun sequence data of Parys Mountain samples suggests an extensive genetic exchange between Mia14 and hosts in situ. Within the subset of organisms with high-quality genomic assemblies representing the âDPANNâ superphylum, the Mia14 lineage has had the largest gene flux, with dozens of genes gained that are implicated in the host interaction.</jats:p
Microbial ÎČ-glucosidases from cow rumen metagenome enhance the saccharification of lignocellulose in combination with commercial cellulase cocktail
BACKGROUND: A complete saccharification of plant polymers is the critical step in the efficient production of bio-alcohols. Beta-glucosidases acting in the degradation of intermediate gluco-oligosaccharides produced by cellulases limit the yield of the final product. RESULTS: In the present work, we have identified and then successfully cloned, expressed, purified and characterised 4 highly active beta-glucosidases from fibre-adherent microbial community from the cow rumen. The enzymes were most active at temperatures 45â55°C and pH 4.0-7.0 and exhibited high affinity and activity towards synthetic substrates such as p-nitrophenyl-beta-D-glucopyranoside (pNPbetaG) and pNP-beta-cellobiose, as well as to natural cello-oligosaccharides ranging from cellobiose to cellopentaose. The apparent capability of the most active beta-glucosidase, herein named LAB25g2, was tested for its ability to improve, at low dosage (31.25 units g(-1) dry biomass, using pNPbetaG as substrate), the hydrolysis of pre-treated corn stover (dry matter content of 20%; 350 g glucan kg(-1) dry biomass) in combination with a beta-glucosidase-deficient commercial Trichoderma reseei cellulase cocktail (5 units g(-1) dry biomass in the basis of pNPbetaG). LAB25g2 increased the final hydrolysis yield by a factor of 20% (44.5 ± 1.7% vs. 34.5 ± 1.5% in control conditions) after 96â120 h as compared to control reactions in its absence or in the presence of other commercial beta-glucosidase preparations. The high stability (half-life higher than 5 days at 50°C and pH 5.2) and 2â38000 fold higher (as compared with reported beta-glucosidases) activity towards cello-oligosaccharides may account for its performance in supplementation assays. CONCLUSIONS: The results suggest that beta-glucosidases from yet uncultured bacteria from animal digestomes may be of a potential interest for biotechnological processes related to the effective bio-ethanol production in combination with low dosage of commercial cellulases
Identification and characterization of carboxyl esterases of gill chamber-associated microbiota in the deep-sea shrimp rimicaris exoculata by using functional metagenomics
The shrimp Rimicaris exoculata dominates the fauna in deep-sea hydrothermal vent sites along the Mid-Atlantic Ridge (depth,
2,320 m). Here, we identified and biochemically characterized three carboxyl esterases from microbial communities inhabiting
the R. exoculata gill that were isolated by naive screens of a gill chamber metagenomic library. These proteins exhibit low to
moderate identity to known esterase sequences (<52%) and to each other (11.9 to 63.7%) and appear to have originated from
unknown species or from genera of Proteobacteria related to Thiothrix/Leucothrix (MGS-RG1/RG2) and to the Rhodobacteraceae
group (MGS-RG3). A library of 131 esters and 31 additional esterase/lipase preparations was used to evaluate the activity
profiles of these enzymes. All 3 of these enzymes had greater esterase than lipase activity and exhibited specific activities with
ester substrates (<356Umg 1) in the range of similar enzymes. MGS-RG3 was inhibited by salts and pressure and had a low
optimal temperature (30°C), and its substrate profile clustered within a group of low-activity and substrate-restricted marine
enzymes. In contrast, MGS-RG1 and MGS-RG2 were most active at 45 to 50°C and were salt activated and barotolerant. They
also exhibited wider substrate profiles that were close to those of highly active promiscuous enzymes from a marine hydrothermal
vent (MGS-RG2) and from a cold brackish lake (MGS-RG1). The data presented are discussed in the context of promoting
the examination of enzyme activities of taxa found in habitats that have been neglected for enzyme prospecting; the enzymes
found in these taxa may reflect distinct habitat-specific adaptations and may constitute new sources of rare reaction specificities.The European
Community project MAMBA (FP7-KBBE-2008-226977), grant BIO2011-25012 from the Spanish Ministry of the
Economy and Competitiveness (formerly MICINN). P.N.G. and O.V.G.
were supported by EU FP7 project MICROB3 (FP7-OCEAN.2011
287589). This work received support from the Government of Canada
through Genome Canada and the Ontario Genomics Institute (grant
2009-OGI-ABC-1405 to A.F.Y. and A.S.) and from the U.S. National Institutes
of Health (grants GM074942 and GM094585 to A.S. through the
Midwest Center for Structural Genomics).http://aem.asm.orgam201
Genome sequence and functional genomic analysis of the oil-degrading bacterium Oleispira antarctica
M.K. and P.N.G. designed the work; T.N.C. performed physiological studies; M.K., M.F.,
Y.A.-R., A.B., N.L.-C., M.E.G., O.R.K., T.Y.N., S.K., I.L., O.V.G., M.M.Y. R.R. and P.N.G.
were associated with genome annotation; H.J.H. performed lipids and FAME analysis;
M.F., M-l.F., S.J., S.C. and J.P.A performed chaperonin anti-proteome analysis; A.-x. S.,
O.K., O.E., P.A.P., P.S. and Y.K. were associated with structural proteomics; A.T. and R.F.
were associated with functional proteomics; H.L. performed electron microscopy; R.D.
performed real-time PCR; M.M.-G. and M.F. performed DIGE proteome analysis;
M.G. was involved in siderophore production; O.N.R. performed genomic islandsâ
analysis; H.T. performed storage lipid compoundsâ analysis; P.N.G. coordinated
manuscript writing.Accession Codes: The genome sequence of Oleispira antarctica RB-8 has been deposited
in GenBank under accession core FO203512. Protein structures have deposited in PDB
under accession codes 3QVM (a/b hydrolase, OLEAN_C08020), 3QVQ (phosphodiesterase,
OLEAN_C20330), 3M16 (transaldolase, OLEAN_C18160), 3LQY (isochorismatase,
OLEAN_C07660), 3LNP (amidohydrolase, OLEAN_C13880), 3V77/3L53 (fumarylacetoacetate isomerase/hydrolase, OLEAN_C35840), 3VCR/3LAB
(2-keto-3-deoxy-6-phosphogluconate aldolase, OLEAN_C25130), 3IRU (phoshonoacetaldehyde
hydrolase, OLEAN_C33610), 3I4Q (inorganic pyrophosphatase,
OLEAN_C30460), 3LMB (protein with unknown function, OLEAN_C10530).Ubiquitous bacteria from the genus Oleispira drive oil degradation in the largest environment
on Earth, the cold and deep sea. Here we report the genome sequence of Oleispira antarctica
and show that compared with Alcanivorax borkumensisâthe paradigm of mesophilic
hydrocarbonoclastic bacteriaâO. antarctica has a larger genome that has witnessed massive
gene-transfer events. We identify an array of alkane monooxygenases, osmoprotectants,
siderophores and micronutrient-scavenging pathways. We also show that at low temperatures,
the main protein-folding machine Cpn60 functions as a single heptameric barrel that
uses larger proteins as substrates compared with the classical double-barrel structure
observed at higher temperatures. With 11 protein crystal structures, we further report the
largest set of structures from one psychrotolerant organism. The most common structural
feature is an increased content of surface-exposed negatively charged residues compared to
their mesophilic counterparts. Our findings are relevant in the context of microbial
cold-adaptation mechanisms and the development of strategies for oil-spill mitigation in cold
environments.We acknowledge the funding from the EU Framework Program 7 to support Projects
MAMBA (226977), ULIXES (266473), MAGIC PAH (245226) and MICROB3 (287589)
This work received the support of the Government of Canada through Genome Canada
and the Ontario Genomics Institute (grant 2009-OGI-ABC-1405 to A.F.Y. and A.S.), and
the U.S. Government National Institutes of Health (grants GM074942 and GM094585
(to A.S. through Midwest Center for Structural Genomics). The study was supported by
the Max Planck Society and the Deutsche Forschungsgemeinschaft through project KU
2679/2-1 and BU 890/21-1. We thank the sequencing team of the AG Reinhardt for
technical assistance and Alfred Beck for computational support. The skilful work of
electron microscopic sample preparation by Mrs. Ingeborg Kristen (Dept. VAM, HZI
Braunschweig) is gratefully acknowledged. Authors thank Professor Ken Timmis for his
critical reading the manuscript and useful comments.http://www.nature.com/naturecommunicationsam201
HPLC-MS/MS Trachypus boharti Brazil 2
HPLC-MS/MS runs of MeOH extracts from single Trachypus boharti specimen antennae collected in Brazil - second hal
HPLC-MS/MS Philanthus spp. South Africa
HPLC-MS/MS runs of MeOH extracts from single Philanthus specimen antennae collected in South Afric
HPLC-MS/MS Philanthus gibbosus cocoons
HPLC-MS/MS runs of MeOH extracts from single Philanthus gibbosus cocoons reared in German