bluuebunny/crossref_metadata_2025_split huggingface dataset:
Stable Isotope Probing Analysis of Interactions between Ammonia Oxidizers
Authors: Tourna Maria , Freitag Thomas E. , Prosser James I. 
Published: February 2010

ABSTRACT

The response of natural microbial communities to environmental change can be assessed by determining DNA- or RNA-targeted changes in relative abundance of 16S rRNA gene sequences by using fingerprinting techniques such as denaturing gradient gel electrophoresis (DNA-DGGE and RNA-DGGE, respectively) or by stable isotope probing (SIP) of 16S rRNA genes following incubation with a
13
C-labeled substrate (DNA-SIP-DGGE). The sensitivities of these three approaches were compared during batch growth of communities containing two or three
Nitrosospira
pure or enriched cultures with different tolerances to a high ammonia concentration. Cultures were supplied with low, intermediate, or high initial ammonia concentrations and with
13
C-labeled carbon dioxide. DNA-SIP-DGGE provided the most direct evidence for growth and was the most sensitive, with changes in DGGE profiles evident before changes in DNA- and RNA-DGGE profiles and before detectable increases in nitrite and nitrate production. RNA-DGGE provided intermediate sensitivity. In addition, the three molecular methods were used to follow growth of individual strains within communities. In general, changes in relative activities of individual strains within communities could be predicted from monoculture growth characteristics. Ammonia-tolerant
Nitrosospira
cluster 3b strains dominated mixed communities at all ammonia concentrations, and ammonia-sensitive strains were outcompeted at an intermediate ammonia concentration. However, coexistence of ammonia-tolerant and ammonia-sensitive strains occurred at the lowest ammonia concentration, and, under some conditions, strains inhibited at high ammonia in monoculture were active at high ammonia in mixed cultures, where they coexisted with ammonia-tolerant strains. The results therefore demonstrate the sensitivity of SIP for detection of activity of organisms with relatively low yield and low activity and its ability to follow changes in the structure of interacting microbial communities.
The Genome Sequence ofPsychrobacter arcticus273-4, a Psychroactive Siberian Permafrost Bacterium, Reveals Mechanisms for Adaptation to Low-Temperature Growth
Authors: Ayala-del-Río Héctor L. , Chain Patrick S. , Grzymski Joseph J. , Ponder Monica A. , Ivanova Natalia , Bergholz Peter W. , Di Bartolo Genevive , Hauser Loren , Land Miriam , Bakermans Corien , Rodrigues Debora , Klappenbach Joel , Zarka Dan , Larimer Frank , Richardson Paul , Murray Alison , Thomashow Michael , Tiedje James M. 
Published: February 2010

ABSTRACTPsychrobacter arcticusstrain 273-4, which grows at temperatures as low as −10°C, is the first cold-adapted bacterium from a terrestrial environment whose genome was sequenced. Analysis of the 2.65-Mb genome suggested that some of the strategies employed byP. arcticus273-4 for survival under cold and stress conditions are changes in membrane composition, synthesis of cold shock proteins, and the use of acetate as an energy source. Comparative genome analysis indicated that in a significant portion of theP. arcticusproteome there is reduced use of the acidic amino acids and proline and arginine, which is consistent with increased protein flexibility at low temperatures. Differential amino acid usage occurred in all gene categories, but it was more common in gene categories essential for cell growth and reproduction, suggesting thatP. arcticusevolved to grow at low temperatures. Amino acid adaptations and the gene content likely evolved in response to the long-term freezing temperatures (−10°C to −12°C) of the Kolyma (Siberia) permafrost soil from which this strain was isolated. Intracellular water likely does not freeze at thesein situtemperatures, which allowsP. arcticusto live at subzero temperatures.
New Vector System for Random, Single-Step Integration of Multiple Copies of DNA into the
Rhodococcus
Genome
Authors: Sallam Khalid Ibrahim , Tamura Noriko , Imoto Noriko , Tamura Tomohiro 
Published: February 2010

ABSTRACT

We designed a new vector system for creating a random mutant library with multiple integrations of DNA fragments into the
Rhodococcus
genome in a single step. For this, we cotransformed two vectors into
Rhodococcus
by electroporation: pTip-
istAB
-
sacB
regulates the expression of the transposase (IstA) and its helper protein (IstB) under the influence of a thiostrepton-inducible promoter, and pRTSK-
sacB
provides the transposable-marker DNA. Both are multicopy vectors that are stable in the host cells; transposition of the transposable-marker DNA occurs only after the induction of IstA/IstB expression. With the addition of thiostrepton, all cultured cells harboring the two vectors, irrespective of the volume, can be mutated by random insertion of the transposable-marker DNA into their genome. Among the generated mutants examined, 30% showed multiple (two to five) insertion copies. The multiple integrated DNA copies were stable in the genome for more than 80 generations of serial growth without the addition of any selective antibiotics. This system can also be used for integrating various copy numbers of stably maintained protein expression cassettes in the host cell genome to modulate the expression level of biologically active recombinant proteins. We successfully applied this system to integrate multiple copies of expression cassettes for proline iminopeptidase and vitamin D
3
hydroxylase into the
Rhodococcus
genome and verified that the clones containing double or multiple copies of the integrated cassettes produced higher levels and showed higher enzymatic activities of the target protein than clones with only a single copy of integration.
Effects of Simulated Mars Conditions on the Survival and Growth of
Escherichia coli
and
Serratia liquefaciens
Authors: Berry Bonnie J. , Jenkins David G. , Schuerger Andrew C. 
Published: February 2010

ABSTRACT

Escherichia coli
and
Serratia liquefaciens
, two bacterial spacecraft contaminants known to replicate under low atmospheric pressures of 2.5 kPa, were tested for growth and survival under simulated Mars conditions. Environmental stresses of high salinity, low temperature, and low pressure were screened alone and in combination for effects on bacterial survival and replication, and then cells were tested in Mars analog soils under simulated Mars conditions. Survival and replication of
E. coli
and
S. liquefaciens
cells in liquid medium were evaluated for 7 days under low temperatures (5, 10, 20, or 30°C) with increasing concentrations (0, 5, 10, or 20%) of three salts (MgCl
2
, MgSO
4
, NaCl) reported to be present on the surface of Mars. Moderate to high growth rates were observed for
E. coli
and
S. liquefaciens
at 30 or 20°C and in solutions with 0 or 5% salts. In contrast, cell densities of both species generally did not increase above initial inoculum levels under the highest salt concentrations (10 and 20%) and the four temperatures tested, with the exception that moderately higher cell densities were observed for both species at 10% MgSO
4
maintained at 20 or 30°C. Growth rates of
E. coli
and
S. liquefaciens
in low salt concentrations were robust under all pressures (2.5, 10, or 101.3 kPa), exhibiting a general increase of up to 2.5 orders of magnitude above the initial inoculum levels of the assays. Vegetative
E. coli
cells were maintained in a Mars analog soil for 7 days under simulated Mars conditions that included temperatures between 20 and −50°C for a day/night diurnal period, UVC irradiation (200 to 280 nm) at 3.6 W m
−2
for daytime operations (8 h), pressures held at a constant 0.71 kPa, and a gas composition that included the top five gases found in the martian atmosphere. Cell densities of
E. coli
failed to increase under simulated Mars conditions, and survival was reduced 1 to 2 orders of magnitude by the interactive effects of desiccation, UV irradiation, high salinity, and low pressure (in decreasing order of importance). Results suggest that
E. coli
may be able to survive, but not grow, in surficial soils on Mars.
Regulation of Morphogenesis and Biocontrol Properties inTrichoderma virensby a VELVET Protein, Vel1
Authors: Mukherjee Prasun K. , Kenerley Charles M. 
Published: February 2010

ABSTRACTMycoparasitic strains ofTrichodermaare applied as commercial biofungicides for control of soilborne plant pathogens. Although the majority of commercial biofungicides areTrichodermabased, chemical pesticides, which are ecological and environmental hazards, still dominate the market. This is because biofungicides are not as effective or consistent as chemical fungicides. Efforts to improve these products have been limited by a lack of understanding of the genetic regulation of biocontrol activities. In this study, using gene knockout and complementation, we identified the VELVET protein Vel1 as a key regulator of biocontrol, as well as morphogenetic traits, inTrichoderma virens, a commercial biocontrol agent. Mutants with mutations invel1were defective in secondary metabolism (antibiosis), mycoparasitism, and biocontrol efficacy. In nutrient-rich media they also lacked two types of spores important for survival and development of formulation products: conidia (on agar) and chlamydospores (in liquid shake cultures). These findings provide an opportunity for genetic enhancement of biocontrol and industrial strains ofTrichoderma, since Vel1 is very highly conserved across threeTrichodermaspecies.
Census of the Viral Metagenome within an Activated Sludge Microbial Assemblage
Authors: Parsley Larissa C. , Consuegra Erin J. , Thomas Stephen J. , Bhavsar Jaysheel , Land Andrew M. , Bhuiyan Nadia N. , Mazher Mustafa A. , Waters Robert J. , Wommack K. Eric , Harper Willie F. , Liles Mark R. 
Published: February 2010

ABSTRACT
The viral metagenome within an activated sludge microbial assemblage was sampled using culture-dependent and culture-independent methods and compared to the diversity of activated sludge bacterial taxa. A total of 70 unique cultured bacterial isolates, 24 cultured bacteriophages, 829 bacterial metagenomic clones of 16S rRNA genes, and 1,161 viral metagenomic clones were subjected to a phylogenetic analysis.
Distribution of Virulence Factors and Molecular Fingerprinting of
Aeromonas
Species Isolates from Water and Clinical Samples: Suggestive Evidence of Water-to-Human Transmission
Authors: Khajanchi Bijay K. , Fadl Amin A. , Borchardt Mark A. , Berg Richard L. , Horneman Amy J. , Stemper Mary E. , Joseph Sam W. , Moyer Nelson P. , Sha Jian , Chopra Ashok K. 
Published: February 2010

ABSTRACT

A total of 227 isolates of
Aeromonas
obtained from different geographical locations in the United States and different parts of the world, including 28 reference strains, were analyzed to determine the presence of various virulence factors. These isolates were also fingerprinted using biochemical identification and pulse-field gel electrophoresis (PFGE). Of these 227 isolates, 199 that were collected from water and clinical samples belonged to three major groups or complexes, namely, the
A. hydrophila
group, the
A. caviae-A. media
group, and the
A. veronii-A. sobria
group, based on biochemical profiles, and they had various pulsotypes. When virulence factor activities were examined,
Aeromonas
isolates obtained from clinical sources had higher cytotoxic activities than isolates obtained from water sources for all three
Aeromonas
species groups. Likewise, the production of quorum-sensing signaling molecules, such as
N
-acyl homoserine lactone, was greater in clinical isolates than in isolates from water for the
A. caviae-A. media
and
A. hydrophila
groups. Based on colony blot DNA hybridization, the heat-labile cytotonic enterotoxin gene and the DNA adenosine methyltransferase gene were more prevalent in clinical isolates than in water isolates for all three
Aeromonas
groups. Using colony blot DNA hybridization and PFGE, we obtained three sets of water and clinical isolates that had the same virulence signature and had indistinguishable PFGE patterns. In addition, all of these isolates belonged to the
A. caviae-A. media
group. The findings of the present study provide the first suggestive evidence of successful colonization and infection by particular strains of certain
Aeromonas
species after transmission from water to humans.
Isolation of Novel Extreme-Tolerant Cyanobacteria from a Rock-Dwelling Microbial Community by Using Exposure to Low Earth Orbit
Authors: Olsson-Francis Karen , de la Torre Rosa , Cockell Charles S. 
Published: February 2010

ABSTRACT

Many cyanobacteria are known to tolerate environmental extremes. Motivated by an interest in selecting cyanobacteria for applications in space, we launched rocks from a limestone cliff in Beer, Devon, United Kingdom, containing an epilithic and endolithic rock-dwelling community of cyanobacteria into low Earth orbit (LEO) at a height of approximately 300 kilometers. The community was exposed for 10 days to isolate cyanobacteria that can survive exposure to the extreme radiation and desiccating conditions associated with space. Culture-independent (16S rRNA) and culture-dependent methods showed that the cyanobacterial community was composed of
Pleurocapsales
,
Oscillatoriales
, and
Chroococcales
. A single cyanobacterium, a previously uncharacterized extremophile, was isolated after exposure to LEO. We were able to isolate the cyanobacterium from the limestone cliff after exposing the rock-dwelling community to desiccation and vacuum (0.7 × 10
−3
kPa) in the laboratory. The ability of the organism to survive the conditions in space may be linked to the formation of dense colonies. These experiments show how extreme environmental conditions, including space, can be used to select for novel microorganisms. Furthermore, it improves our knowledge of environmental tolerances of extremophilic rock-dwelling cyanobacteria.
Requirements for Construction of a Functional Hybrid Complex of Photosystem I and [NiFe]-Hydrogenase
Authors: Schwarze Alexander , Kopczak Marta J. , Rögner Matthias , Lenz Oliver 
Published: February 2010

ABSTRACT

The development of cellular systems in which the enzyme hydrogenase is efficiently coupled to the oxygenic photosynthesis apparatus represents an attractive avenue to produce H
2
sustainably from light and water. Here we describe the molecular design of the individual components required for the direct coupling of the O
2
-tolerant membrane-bound hydrogenase (MBH) from
Ralstonia eutropha
H16 to the acceptor site of photosystem I (PS I) from
Synechocystis
sp. PCC 6803. By genetic engineering, the peripheral subunit PsaE of PS I was fused to the MBH, and the resulting hybrid protein was purified from
R. eutropha
to apparent homogeneity via two independent affinity chromatographical steps. The catalytically active MBH-PsaE (MBH
PsaE
) hybrid protein could be isolated only from the cytoplasmic fraction. This was surprising, since the MBH is a substrate of the twin-arginine translocation system and was expected to reside in the periplasm. We conclude that the attachment of the additional PsaE domain to the small, electron-transferring subunit of the MBH completely abolished the export competence of the protein. Activity measurements revealed that the H
2
production capacity of the purified MBH
PsaE
fusion protein was very similar to that of wild-type MBH. In order to analyze the specific interaction of MBH
PsaE
with PS I, His-tagged PS I lacking the PsaE subunit was purified via Ni-nitrilotriacetic acid affinity and subsequent hydrophobic interaction chromatography. Formation of PS I-hydrogenase supercomplexes was demonstrated by blue native gel electrophoresis. The results indicate a vital prerequisite for the quantitative analysis of the MBH
PsaE
-PS I complex formation and its light-driven H
2
production capacity by means of spectroelectrochemistry.
Sequence-Based Analysis of Secondary-Metabolite Biosynthesis in Marine Actinobacteria
Authors: Gontang Erin A. , Gaudêncio Susana P. , Fenical William , Jensen Paul R. 
Published: February 2010

ABSTRACT

A diverse collection of 60 marine-sediment-derived
Actinobacteria
representing 52 operational taxonomic units was screened by PCR for genes associated with secondary-metabolite biosynthesis. Three primer sets were employed to specifically target adenylation domains associated with nonribosomal peptide synthetases (NRPSs) and ketosynthase (KS) domains associated with type I modular, iterative, hybrid, and enediyne polyketide synthases (PKSs). In total, two-thirds of the strains yielded a sequence-verified PCR product for at least one of these biosynthetic types. Genes associated with enediyne biosynthesis were detected in only two genera, while 88% of the ketosynthase sequences shared greatest homology with modular PKSs. Positive strains included representatives of families not traditionally associated with secondary-metabolite production, including the
Corynebacteriaceae
,
Gordoniaceae, Intrasporangiaceae
, and
Micrococcaceae
. In four of five cases where phylogenetic analyses of KS sequences revealed close evolutionary relationships to genes associated with experimentally characterized biosynthetic pathways, secondary-metabolite production was accurately predicted. Sequence clustering patterns were used to provide an estimate of PKS pathway diversity and to assess the biosynthetic richness of individual strains. The detection of highly similar KS sequences in distantly related strains provided evidence of horizontal gene transfer, while control experiments designed to amplify KS sequences from
Salinispora arenicola
strain CNS-205, for which a genome sequence is available, led to the detection of 70% of the targeted PKS pathways. The results provide a bioinformatic assessment of secondary-metabolite biosynthetic potential that can be applied in the absence of fully assembled pathways or genome sequences. The rapid identification of strains that possess the greatest potential to produce new secondary metabolites along with those that produce known compounds can be used to improve the process of natural-product discovery by providing a method to prioritize strains for fermentation studies and chemical analysis.
Laccase and Its Role in Production of Extracellular Reactive Oxygen Species during Wood Decay by the Brown Rot Basidiomycete
Postia placenta
Authors: Wei Dongsheng , Houtman Carl J. , Kapich Alexander N. , Hunt Christopher G. , Cullen Daniel , Hammel Kenneth E. 
Published: February 2010

ABSTRACT

Brown rot basidiomycetes initiate wood decay by producing extracellular reactive oxygen species that depolymerize the structural polysaccharides of lignocellulose. Secreted fungal hydroquinones are considered one contributor because they have been shown to reduce Fe
3+
, thus generating perhydroxyl radicals and Fe
2+
, which subsequently react further to produce biodegradative hydroxyl radicals. However, many brown rot fungi also secrete high levels of oxalate, which chelates Fe
3+
tightly, making it unreactive with hydroquinones. For hydroquinone-driven hydroxyl radical production to contribute in this environment, an alternative mechanism to oxidize hydroquinones is required. We show here that aspen wood undergoing decay by the oxalate producer
Postia placenta
contained both 2,5-dimethoxyhydroquinone and laccase activity. Mass spectrometric analysis of proteins extracted from the wood identified a putative laccase (Joint Genome Institute
P. placenta
protein identification number 111314), and heterologous expression of the corresponding gene confirmed this assignment. Ultrafiltration experiments with liquid pressed from the biodegrading wood showed that a high-molecular-weight component was required for it to oxidize 2,5-dimethoxyhydroquinone rapidly and that this component was replaceable by
P. placenta
laccase. The purified laccase oxidized 2,5-dimethoxyhydroquinone with a second-order rate constant near 10
4
M
−1
s
−1
, and measurements of the H
2
O
2
produced indicated that approximately one perhydroxyl radical was generated per hydroquinone supplied. Using these values and a previously developed computer model, we estimate that the quantity of reactive oxygen species produced by
P. placenta
laccase in wood is large enough that it likely contributes to incipient decay.
Changes in Denitrifier Abundance, Denitrification Gene mRNA Levels, Nitrous Oxide Emissions, and Denitrification in Anoxic Soil Microcosms Amended with Glucose and Plant Residues
Authors: Henderson Sherri L. , Dandie Catherine E. , Patten Cheryl L. , Zebarth Bernie J. , Burton David L. , Trevors Jack T. , Goyer Claudia 
Published: February 2010

ABSTRACTIn agricultural cropping systems, crop residues are sources of organic carbon (C), an important factor influencing denitrification. The effects of red clover, soybean, and barley plant residues and of glucose on denitrifier abundance, denitrification gene mRNA levels, nitrous oxide (N2O) emissions, and denitrification rates were quantified in anoxic soil microcosms for 72 h.nosZgene abundances and mRNA levels significantly increased in response to all organic carbon treatments over time. In contrast, the abundance and mRNA levels ofPseudomonas mandeliiand closely related species (nirSP) increased only in glucose-amended soil: thenirSPguild abundance increased 5-fold over the 72-h incubation period (P< 0.001), while the mRNA level significantly increased more than 15-fold at 12 h (P< 0.001) and then subsequently decreased. ThenosZgene abundance was greater in plant residue-amended soil than in glucose-amended soil. Although plant residue carbon-to-nitrogen (C:N) ratios varied from 15:1 to 30:1,nosZgene and mRNA levels were not significantly different among plant residue treatments, with an average of 3.5 × 107gene copies and 6.9 × 107transcripts g−1dry soil. Cumulative N2O emissions and denitrification rates increased over 72 h in both glucose- and plant-tissue-C-treated soil. ThenirSPandnosZcommunities responded differently to glucose and plant residue amendments. However, the targeted denitrifier communities responded similarly to the different plant residues under the conditions tested despite changes in the quality of organic C and different C:N ratios.