bluuebunny/crossref_metadata_2025_split huggingface dataset:
Enhanced Growth of Acidovorax sp. Strain 2AN during Nitrate-Dependent Fe(II) Oxidation in Batch and Continuous-Flow Systems
Authors: Chakraborty Anirban , Roden Eric E. , Schieber Jürgen , Picardal Flynn 
Published: October 2011

ABSTRACT

Microbial nitrate-dependent, Fe(II) oxidation (NDFO) is a ubiquitous biogeochemical process in anoxic sediments. Since most microorganisms that can oxidize Fe(II) with nitrate require an additional organic substrate for growth or sustained Fe(II) oxidation, the energetic benefits of NDFO are unclear. The process may also be self-limiting in batch cultures due to formation of Fe-oxide cell encrustations. We hypothesized that NDFO provides energetic benefits via a mixotrophic physiology in environments where cells encounter very low substrate concentrations, thereby minimizing cell encrustations.
Acidovorax
sp. strain 2AN was incubated in anoxic batch reactors in a defined medium containing 5 to 6 mM NO
3
−
, 8 to 9 mM Fe
2+
, and 1.5 mM acetate. Almost 90% of the Fe(II) was oxidized within 7 days with concomitant reduction of nitrate and complete consumption of acetate. Batch-grown cells became heavily encrusted with Fe(III) oxyhydroxides, lost motility, and formed aggregates. Encrusted cells could neither oxidize more Fe(II) nor utilize further acetate additions. In similar experiments with chelated iron (Fe(II)-EDTA), encrusted cells were not produced, and further additions of acetate and Fe(II)-EDTA could be oxidized. Experiments using a novel, continuous-flow culture system with low concentrations of substrate, e.g., 100 μM NO
3
−
, 20 μM acetate, and 50 to 250 μM Fe
2+
, showed that the growth yield of
Acidovorax
sp. strain 2AN was always greater in the presence of Fe(II) than in its absence, and electron microscopy showed that encrustation was minimized. Our results provide evidence that, under environmentally relevant concentrations of substrates, NDFO can enhance growth without the formation of growth-limiting cell encrustations.
Nitrate-Dependent N
2
O Emission from Intact Soybean Nodules via Denitrification by Bradyrhizobium japonicum Bacteroids
Authors: Hirayama Junta , Eda Shima , Mitsui Hisayuki , Minamisawa Kiwamu 
Published: October 2011

ABSTRACT

In the presence of nitrate, N
2
O emission increased markedly from soybean roots inoculated with
nosZ
mutant of
Bradyrhizobium japonicum
, but not from soybean roots inoculated with a
napA nosZ
double mutant, indicating that
B. japonicum
bacteroids in soybean nodules are able to convert the exogenously supplied nitrate into N
2
O via a denitrification pathway.
Global Genome Transcription Profiling of Bifidobacterium bifidum PRL2010 under
In Vitro
Conditions and Identification of Reference Genes for Quantitative Real-Time PCR
Authors: Turroni Francesca , Foroni Elena , Montanini Barbara , Viappiani Alice , Strati Francesco , Duranti Sabrina , Ferrarini Alberto , Delledonne Massimo , van Sinderen Douwe , Ventura Marco 
Published: October 2011

ABSTRACT

Bifidobacteria have attracted significant scientific attention due to their perceived role as health-promoting microorganisms, although the genetics of the bacterial group is still underexplored. In this study, we investigated the transcriptome of
Bifidobacterium bifidum
PRL2010 during
in vitro
growth by microarray technology. When
B. bifidum
PRL2010 was grown in liquid broth, 425 of the 1,644 PRL2010 genes represented on the array were expressed in at least one of the three investigated growth phases, i.e., the lag, exponential, and stationary phases. These transcriptional analyses identified a core
in vitro
transcriptome encompassing 150 genes that are expressed in all phases. A proportion of these genes were further investigated as potential reference genes by quantitative real-time reverse transcription-PCR (qRT-PCR) assays. Their expression stability was evaluated under different growth conditions, which included cultivation on different carbon sources, exposure to environmental stresses (thermal, acidic, and osmotic), and growth phases. Our analyses validated six reference genes suitable for normalizing mRNA expression levels in qRT-PCR experiments applied to bifidobacteria.
Connection between Multimetal(loid) Methylation in Methanoarchaea and Central Intermediates of Methanogenesis
Authors: Thomas Frank , Diaz-Bone Roland A. , Wuerfel Oliver , Huber Britta , Weidenbach Katrin , Schmitz Ruth A. , Hensel Reinhard 
Published: October 2011

ABSTRACT

In spite of the significant impact of biomethylation on the mobility and toxicity of metals and metalloids in the environment, little is known about the biological formation of these methylated metal(loid) compounds. While element-specific methyltransferases have been isolated for arsenic, the striking versatility of methanoarchaea to methylate numerous metal(loid)s, including rare elements like bismuth, is still not understood. Here, we demonstrate that the same metal(loid)s (arsenic, selenium, antimony, tellurium, and bismuth) that are methylated by
Methanosarcina mazei in vivo
are also methylated by
in vitro
assays with purified recombinant MtaA, a methyltransferase catalyzing the methyl transfer from methylcobalamin [CH
3
Cob(III)] to 2-mercaptoethanesulfonic acid (CoM) in methylotrophic methanogenesis. Detailed studies revealed that cob(I)alamin [Cob(I)], formed by MtaA-catalyzed demethylation of CH
3
Cob(III), is the causative agent for the multimetal(loid) methylation observed. Moreover, Cob(I) is also capable of metal(loid) hydride generation. Global transcriptome profiling of
M. mazei
cultures exposed to bismuth did not reveal induced methyltransferase systems but upregulated regeneration of methanogenic cofactors in the presence of bismuth. Thus, we conclude that the multimetal(loid) methylation
in vivo
is attributed to side reactions of CH
3
Cob(III) with reduced cofactors formed in methanogenesis. The close connection between metal(loid) methylation and methanogenesis explains the general capability of methanoarchaea to methylate metal(loid)s.
Lactate Oxidation Coupled to Iron or Electrode Reduction by Geobacter sulfurreducens PCA
Authors: Call Douglas F. , Logan Bruce E. 
Published: October 2011

ABSTRACT

Geobacter sulfurreducens
PCA completely oxidized lactate and reduced iron or an electrode, producing pyruvate and acetate intermediates. Compared to the current produced by
Shewanella oneidensis
MR-1,
G. sulfurreducens
PCA produced 10-times-higher current levels in lactate-fed microbial electrolysis cells. The kinetic and comparative analyses reported here suggest a prominent role of
G. sulfurreducens
strains in metal- and electrode-reducing communities supplied with lactate.
Genetic Background and Mobility of Variants of the Gene
nleA
in Attaching and Effacing Escherichia coli
Authors: Creuzburg Kristina , Heeren Sabine , Lis Claudia M. , Kranz Markus , Hensel Michael , Schmidt Herbert 
Published: October 2011

ABSTRACT

In this study, we characterized the genetic background of various
nleA
variants in 106 Shiga toxin-producing
Escherichia coli
(STEC) and enteropathogenic
Escherichia coli
(EPEC) strains. The flanking regions of eight
nleA
variants were analyzed by DNA sequencing and compared with the corresponding regions of five previously described NleA-encoding prophages. The analyzed
nleA
variants were all located downstream of the DNA region responsible for phage morphogenesis. In particular, the type III effector genes
avrA
,
ospB
,
nleH
, and
nleG
and IS elements were detected in the neighborhood of
nleA
. The structure of the eight analyzed regions flanking
nleA
primarily resembled the corresponding region of the NleA
4795
-encoding prophage BP-4795. Using PCR, the gene order flanking 13
nleA
variants in strains of different serogroups was compared to the respective regions in reference strains. The analyses showed that strains which harbor prophages with conserved flanking regions of a particular
nleA
variant predominantly occurred, and IS elements were additionally detected in these regions. We were able to mobilize
nleA
by transduction in 20% of strains determined, which comprised in particular EPEC strains harboring an
nleA
variant, the gene encoding the protein known as “EspI-like.” Plaque hybridization was used to identify phages that harbor the genes
stx
and
nleA
. However, only two strains harbored variant
nleA
4795
in the genome of an Stx1 prophage.
A Whole-Genome Single Nucleotide Polymorphism-Based Approach To Trace and Identify Outbreaks Linked to a Common Salmonella enterica subsp. enterica Serovar Montevideo Pulsed-Field Gel Electrophoresis Type
Authors: den Bakker Henk C. , Moreno Switt Andrea I. , Cummings Craig A. , Hoelzer Karin , Degoricija Lovorka , Rodriguez-Rivera Lorraine D. , Wright Emily M. , Fang Rixun , Davis Margaret , Root Tim , Schoonmaker-Bopp Dianna , Musser Kimberlee A. , Villamil Elizabeth , Waechter HaeNa , Kornstein Laura , Furtado Manohar R. , Wiedmann Martin 
Published: October 2011

ABSTRACT

In this study, we report a whole-genome single nucleotide polymorphism (SNP)-based evolutionary approach to study the epidemiology of a multistate outbreak of
Salmonella enterica
subsp.
enterica
serovar Montevideo. This outbreak included 272 cases that occurred in 44 states between July 2009 and April 2010. A case-control study linked the consumption of salami made with contaminated black and red pepper to the outbreak. We sequenced, on the SOLiD System, 47 isolates with XbaI PFGE pattern JIXX01.0011, a common pulsed-field gel electrophoresis (PFGE) pattern associated with isolates from the outbreak. These isolates represented 20 isolates collected from human sources during the period of the outbreak and 27 control isolates collected from human, food, animal, and environmental sources before the outbreak. Based on 253 high-confidence SNPs, we were able to reconstruct a tip-dated molecular clock phylogeny of the isolates and to assign four human isolates to the actual outbreak. We developed an SNP typing assay to rapidly discriminate between outbreak-related cases and non-outbreak-related cases and tested this assay on an extended panel of 112 isolates. These results suggest that only a very small percentage of the human isolates with the outbreak PFGE pattern and obtained during the outbreak period could be attributed to the actual pepper-related outbreak (20%), while the majority (80%) of the putative cases represented background cases. This study demonstrates that next-generation-based SNP typing provides the resolution and accuracy needed for outbreak investigations of food-borne pathogens that cannot be distinguished by currently used subtyping methods.
Novel Shuttle Markers for Nuclear Transformation of the Green Alga Chlamydomonas reinhardtii
Authors: Meslet-Cladière Laurence , Vallon Olivier 
Published: October 2011

ABSTRACT

The green alga
Chlamydomonas reinhardtii
today is a premier model organism for the study of green algae and plants. Yet the efficient engineering of its nuclear genome requires development of new antibiotic resistance markers. We have recoded, based on codon usage in the nuclear genome, the AadA marker that has been used previously for chloroplast transformation. The recoded AadA gene, placed under the control of the
HSP70A-RBCS2
hybrid promoter and preceded by the RbcS2 chloroplast-targeting peptide, can be integrated into the nuclear genome by electroporation, conferring resistance to spectinomycin and streptomycin. Transformation efficiency is markedly increased when vector sequences are completely eliminated from the transforming DNA. Antibiotic resistance is stable for several months in the absence of selection pressure. Shuttle markers allowing selection in both
Chlamydomonas
and
Escherichia coli
would also be a useful asset. By placing an artificial bacterial promoter and Shine-Dalgarno sequence in frame within the AadA coding sequence, we generated such a shuttle marker. To our surprise, we found that the classical AphVIII construct already functions as a shuttle marker. Finally, we developed a method to introduce the AadA and AphVIII markers into the vector part of the bacterial artificial chromosomes (BACs) of the
Chlamydomonas
genomic DNA library. Our aim was to facilitate complementation studies whenever the test gene cannot be selected for directly. After transformation of a
petC
mutant with a modified BAC carrying the AphVIII marker along with the
PETC
gene in the insert, almost half of the paromomycin-resistant transformants obtained showed restoration of phototrophy, indicating successful integration of the unselected test gene. With AadA, cotransformation was also observed, but with a lower efficiency.
The C2 Domain Protein Cts1 Functions in the Calcineurin Signaling Circuit during High-Temperature Stress Responses in Cryptococcus neoformans
Authors: Aboobakar Eanas F. , Wang Xuying , Heitman Joseph , Kozubowski Lukasz 
Published: October 2011

ABSTRACTCalcineurin is a conserved calcium/calmodulin-dependent serine/threonine-specific protein phosphatase that acts in cell stress responses. Calcineurin is essential for growth at 37°C and for virulence of the human fungal pathogenCryptococcus neoformans, but its substrates remain unknown. The C2 domain-containing, phospholipid-binding protein Cts1 was previously identified as a multicopy suppressor of a calcineurin mutation inC. neoformans. Here we further characterize the function of Cts1 and the links between Cts1 and calcineurin. GFP-Cts1 localizes to cytoplasmic puncta and colocalizes with the endosomal marker FM4-64. Thects1Δ mutant shows a distinct FM4-64 staining pattern, suggesting involvement of Cts1 in endocytic trafficking. In large budded cells, GFP-Cts1 localizes transiently at the mother bud neck, as a single ring that undergoes contraction. mCherry-Cts1 colocalizes with the GFP-tagged calcineurin catalytic subunit Cna1 at sites of mRNA processing at 37°C, suggesting that Cts1 and calcineurin function coordinately during thermal stress. GFP-Cts1 exhibits slower electrophoretic mobility for cells grown at 37°C than for cells grown at 24°C, and the shift to a higher molecular weight is more pronounced in the presence of the calcineurin inhibitor FK506.In vitrotreatment with calf intestinal alkaline phosphatase (CIP) restores faster electrophoretic mobility to GFP-Cts1, suggesting that Cts1 is phosphorylated at 37°C and may be dephosphorylated in a calcineurin-dependent manner. mCherry-Cts1 also coimmunoprecipitates with GFP-Cna1, with greater complex formation at 37°C than at 24°C. Taken together, these findings support potential roles for Cts1 in endocytic trafficking, mRNA processing, and cytokinesis and suggest that Cts1 is a substrate of calcineurin during high-temperature stress responses.
Reg1 Protein Regulates Phosphorylation of All Three Snf1 Isoforms but Preferentially Associates with the Gal83 Isoform
Authors: Zhang Yuxun , McCartney Rhonda R. , Chandrashekarappa Dakshayini G. , Mangat Simmanjeet , Schmidt Martin C. 
Published: October 2011

ABSTRACT
The phosphorylation status of the Snf1 activation loop threonine is determined by changes in the rate of its dephosphorylation, catalyzed by the yeast PP1 phosphatase Glc7 in complex with the Reg1 protein. Previous studies have shown that Reg1 can associate with both Snf1 and Glc7, suggesting substrate binding as a mechanism for Reg1-mediated targeting of Glc7. In this study, the association of Reg1 with the three Snf1 isoforms was measured by two-hybrid analysis and coimmunoprecipitation. We found that Reg1 association with Snf1 occurred almost exclusively with the Gal83 isoform of the Snf1 complex. Nonetheless, Reg1 plays an important role in determining the phosphorylation status of all three Snf1 isoforms. We found that the rate of dephosphorylation for isoforms of Snf1 did not correlate with the amount of associated Reg1 protein. Functional chimeric β subunits containing residues from Gal83 and Sip2 were used to map the residues needed to promote Reg1 association with the N-terminal 150 residues of Gal83. The Gal83 isoform of Snf1 is the only isoform capable of nuclear localization. A Gal83-Sip2 chimera containing the first 150 residues of Gal83 was able to associate with the Reg1 protein but did not localize to the nucleus. Therefore, nuclear localization is not required for Reg1 association. Taken together, these data indicate that the ability of Reg1 to promote the dephosphorylation of Snf1 is not directly related to the strength of its association with the Snf1 complex.
Disulfide Bonding within Components of the Chlamydia Type III Secretion Apparatus Correlates with Development
Authors: Betts-Hampikian H. J. , Fields K. A. 
Published: October 2011

ABSTRACT

Chlamydia
spp. exhibit a unique biphasic developmental cycle whereby infectious elementary bodies (EBs) invade host epithelial cells and differentiate into noninfectious, metabolically active reticulate bodies (RBs). EBs posses a unique outer envelope where rigidity is achieved by disulfide bonding among cysteine-rich envelope-associated proteins. Conversely, these disulfide bonds become reduced in RBs to accommodate vegetative growth, thereby linking the redox status of cysteine-rich envelope proteins with progression of the developmental cycle. We investigated the potential role of disulfide bonding within the chlamydial type III secretion system (T3SS), since activity of this system is also closely linked to development. We focused on structural components of the T3S apparatus that contain an unusually high number of cysteine residues compared to orthologs in other secretion systems. Nonreducing SDS-PAGE revealed that EB-localized apparatus proteins such as CdsF, CdsD, and CdsC form higher-order complexes mediated by disulfide bonding. The most dramatic alterations were detected for the needle protein CdsF. Significantly, disulfide bonding patterns shifted during differentiation of developmental forms and were completely reduced in RBs. Furthermore, at later time points during infection following RB to EB conversion, we found that CdsF is reoxidized into higher-order complexes. Overall, we conclude that the redox status of specific T3SS apparatus proteins is intimately linked to the developmental cycle and constitutes a newly appreciated aspect of functionally significant alterations within proteins of the chlamydial envelope.
Translocators YopB and YopD from Yersinia enterocolitica Form a Multimeric Integral Membrane Complex in Eukaryotic Cell Membranes
Authors: Montagner Caroline , Arquint Christian , Cornelis Guy R. 
Published: October 2011

ABSTRACT

The type III secretion systems are contact-activated secretion systems that allow bacteria to inject effector proteins across eukaryotic cell membranes. The secretion apparatus, called injectisome or needle complex, includes a needle that terminates with a tip structure. The injectisome exports its own distal components, like the needle subunit and the needle tip. Upon contact, it exports two hydrophobic proteins called translocators (YopB and YopD in
Yersinia enterocolitica
) and the effectors. The translocators, assisted by the needle tip, form a pore in the target cell membrane, but the structure of this pore remains elusive. Here, we purified the membranes from infected sheep erythrocytes, and we show that they contain integrated and not simply adherent YopB and YopD. In blue native PAGE, these proteins appeared as a multimeric 500- to 700-kDa complex. This heteropolymeric YopBD complex could be copurified after solubilization in 0.5% dodecyl maltoside but not visualized in the electron microscope. We speculate that this complex may not be stable and rigid but only transient.
The Extended Signal Peptide of the Trimeric Autotransporter EmaA of Aggregatibacter actinomycetemcomitans Modulates Secretion
Authors: Jiang X. , Ruiz T. , Mintz K. P. 
Published: October 2011

ABSTRACT

The extracellular matrix protein adhesin A (EmaA) of the Gram-negative bacterium
Aggregatibacter actinomycetemcomitans
is a fibrillar collagen adhesin belonging to the family of trimeric autotransporters. The protein forms antenna-like structures on the bacterial surface required for collagen adhesion. The 202-kDa protein monomers are proposed to be targeted and translocated across the inner membrane by a long signal peptide composed of 56 amino acids. The predicted signal peptide was functionally active in
Escherichia coli
and
A. actinomycetemcomitans
using truncated PhoA and Aae chimeric proteins, respectively. Mutations in the signal peptide were generated and characterized for PhoA activity in
E. coli
.
A. actinomycetemcomitans
strains expressing EmaA with the identical mutant signal peptides were assessed for cellular localization, surface expression, and collagen binding activity. All of the mutants impaired some aspect of EmaA structure or function. A signal peptide mutant that promoted alkaline phosphatase secretion did not allow any cell surface presentation of EmaA. A second mutant allowed for cell surface exposure but abolished protein function. A third mutant allowed for the normal localization and function of EmaA at 37°C but impaired localization at elevated temperatures. Likewise, replacement of the long EmaA signal peptide with a typical signal peptide also impaired localization above 37°C. The data suggest that the residues of the EmaA signal peptide are required for protein folding or assembly of this collagen adhesin.
Global Transcription Profiles of the Nitrogen Stress Response Resulting in Heterocyst or Hormogonium Development in Nostoc punctiforme
Authors: Christman Harry D. , Campbell Elsie L. , Meeks John C. 
Published: October 2011

ABSTRACT

The filamentous cyanobacterium
Nostoc punctiforme
differentiates from vegetative cells into three distinct cell types, heterocysts, hormogonia, and akinetes, in response to different stimuli. Cultures growing with ammonium can be induced to form hormogonia or heterocysts upon removal of the combined nitrogen. A DNA microarray consisting of 94% of the open reading frames predicted from the 9.059-Mb
N. punctiforme
genome was used to generate a global transcription data set consisting of seven time points over a 24-h period of nitrogen deprivation, which results in heterocyst formation. This data set was compared to a similarly generated data set of nitrogen-starved
N. punctiforme
resulting in hormogonium formation that had previously been published (E. L. Campbell, H. Christman, and J. C. Meeks, J. Bacteriol. 190:7382-7391, 2008). The transition from vegetative cells to either heterocysts or hormogonia resulted in rapid and sustained expression of genes required for utilization of alternate nitrogen sources. Overall, 1,036 and 1,762 genes were found to be differentially transcribed during the heterocyst and hormogonium time courses, respectively, as analyzed with the Bayesian user-friendly software for analyzing time series microarray experiments (BATS). Successive transcription of heterocyst regulatory, structural, and functional genes occurred over the 24 h required to form a functional heterocyst. During hormogonium differentiation, some heterocyst structural and functional genes were upregulated, while the heterocyst master regulator
hetR
was downregulated. There are commonalities in differential expression between cells bound for differentiation into heterocysts or hormogonia, yet the two paths are distinguished by their developmentally specific transcription profiles.