bluuebunny/crossref_metadata_2025_split huggingface dataset:
Diversity of Cultivated Endophytic Bacteria from Sugarcane: Genetic and Biochemical Characterization of
Burkholderia cepacia
Complex Isolates
Authors: Mendes Rodrigo , Pizzirani-Kleiner Aline A. , Araujo Welington L. , Raaijmakers Jos M. 
Published: September 2007

ABSTRACT

Bacteria were isolated from the rhizosphere and from inside the roots and stems of sugarcane plants grown in the field in Brazil. Endophytic bacteria were found in both the roots and the stems of sugarcane plants, with a significantly higher density in the roots. Many of the cultivated endophytic bacteria were shown to produce the plant growth hormone indoleacetic acid, and this trait was more frequently found among bacteria from the stem. 16S rRNA gene sequence analysis revealed that the selected isolates of the endophytic bacterial community of sugarcane belong to the genera of
Burkholderia, Pantoea, Pseudomonas
, and
Microbacterium
. Bacterial isolates belonging to the genus
Burkholderia
were the most predominant among the endophytic bacteria. Many of the
Burkholderia
isolates produced the antifungal metabolite pyrrolnitrin, and all were able to grow at 37°C. Phylogenetic analyses of the 16S rRNA gene and
recA
gene sequences indicated that the endophytic
Burkholderia
isolates from sugarcane are closely related to clinical isolates of the
Burkholderia cepacia
complex and clustered with
B. cenocepacia
(gv. III) isolates from cystic fibrosis patients. These results suggest that isolates of the
B. cepacia
complex are an integral part of the endophytic bacterial community of sugarcane in Brazil and reinforce the hypothesis that plant-associated environments may act as a niche for putative opportunistic human pathogenic bacteria.
Exploration of New Geometries in Cellulosome-Like Chimeras
Authors: Mingardon Florence , Chanal Angélique , Tardif Chantal , Bayer Edward A. , Fierobe Henri-Pierre 
Published: September 2007

ABSTRACTIn this study, novel cellulosome chimeras exhibiting atypical geometries and binding modes, wherein the targeting and proximity functions were directly incorporated as integral parts of the enzyme components, were designed. Two pivotal cellulosomal enzymes (family 48 and 9 cellulases) were thus appended with an efficient cellulose-binding module (CBM) and an optional cohesin and/or dockerin. Compared to the parental enzymes, the chimeric cellulases exhibited improved activity on crystalline cellulose as opposed to their reduced activity on amorphous cellulose. Nevertheless, the various complexes assembled using these engineered enzymes were somewhat less active on crystalline cellulose than the conventional designer cellulosomes containing the parental enzymes. The diminished activity appeared to reflect the number of protein-protein interactions within a given complex, which presumably impeded the mobility of their catalytic modules. The presence of numerous CBMs in a given complex, however, also reduced their performance. Furthermore, a “covalent cellulosome” that combines in a single polypeptide chain a CBM, together with family 48 and family 9 catalytic modules, also exhibited reduced activity. This study also revealed that the cohesin-dockerin interaction may be reversible under specific conditions. Taken together, the data demonstrate that cellulosome components can be used to generate higher-order functional composites and suggest that enzyme mobility is a critical parameter for cellulosome efficiency.
Layered Structure of Bacterial and Archaeal Communities and Their In Situ Activities in Anaerobic Granules
Authors: Satoh Hisashi , Miura Yuki , Tsushima Ikuo , Okabe Satoshi 
Published: September 2007

ABSTRACT

The microbial community structure and spatial distribution of microorganisms and their in situ activities in anaerobic granules were investigated by 16S rRNA gene-based molecular techniques and microsensors for CH
4
, H
2
, pH, and the oxidation-reduction potential (ORP). The 16S rRNA gene-cloning analysis revealed that the clones related to the phyla
Alphaproteobacteria
(detection frequency, 51%),
Firmicutes
(20%),
Chloroflexi
(9%), and
Betaproteobacteria
(8%) dominated the bacterial clone library, and the predominant clones in the archaeal clone library were affiliated with
Methanosaeta
(73%). In situ hybridization with oligonucleotide probes at the phylum level revealed that these microorganisms were numerically abundant in the granule. A layered structure of microorganisms was found in the granule, where
Chloroflexi
and
Betaproteobacteria
were present in the outer shell of the granule,
Firmicutes
were found in the middle layer, and aceticlastic
Archaea
were restricted to the inner layer. Microsensor measurements for CH
4
, H
2
, pH, and ORP revealed that acid and H
2
production occurred in the upper part of the granule, below which H
2
consumption and CH
4
production were detected. Direct comparison of the in situ activity distribution with the spatial distribution of the microorganisms implied that
Chloroflexi
contributed to the degradation of complex organic compounds in the outermost layer, H
2
was produced mainly by
Firmicutes
in the middle layer, and
Methanosaeta
produced CH
4
in the inner layer. We determined the effective diffusion coefficient for H
2
in the anaerobic granules to be 2.66 × 10
−5
cm
2
s
−1
, which was 57% in water.
Caulobacter crescentus
as a Whole-Cell Uranium Biosensor
Authors: Hillson Nathan J. , Hu Ping , Andersen Gary L. , Shapiro Lucy 
Published: September 2007

ABSTRACT

We engineered a strain of the bacterium
Caulobacter crescentus
to fluoresce in the presence of micromolar levels of uranium at ambient temperatures when it is exposed to a hand-held UV lamp. Previous microarray experiments revealed that several
Caulobacter
genes are significantly upregulated in response to uranium but not in response to other heavy metals. We designated one of these genes
urcA
(for
u
ranium
r
esponse in
c
aulobacter). We constructed a reporter that utilizes the
urcA
promoter to produce a UV-excitable green fluorescent protein in the presence of the uranyl cation, a soluble form of uranium. This reporter is specific for uranium and has little cross specificity for nitrate (<400 μM), lead (<150 μM), cadmium (<48 μM), or chromium (<41.6 μM). The uranium reporter construct was effective for discriminating contaminated groundwater samples (4.2 μM uranium) from uncontaminated groundwater samples (<0.1 μM uranium) collected at the Oak Ridge Field Research Center. In contrast to other uranium detection methodologies, the
Caulobacter
reporter strain can provide on-demand usability in the field; it requires minimal sample processing and no equipment other than a hand-held UV lamp, and it may be sprayed directly on soil, groundwater, or industrial surfaces.
LambdaSa1 and LambdaSa2 Prophage Lysins of
Streptococcus agalactiae
Authors: Pritchard David G. , Dong Shengli , Kirk Marion C. , Cartee Robert T. , Baker John R. 
Published: September 2007

ABSTRACT

Putative
N
-acetylmuramyl-
l
-alanine amidase genes from LambdaSa1 and LambdaSa2 prophages of
Streptococcus agalactiae
were cloned and expressed in
Escherichia coli
. The purified enzymes lysed the cell walls of
Streptococcus agalactiae
,
Streptococcus pneumoniae
, and
Staphylococcus aureus
. The peptidoglycan digestion products in the cell wall lysates were not consistent with amidase activity. Instead, the structure of the muropeptide digestion fragments indicated that both the LambdaSa1 and LambdaSa2 lysins exhibited γ-
d
-glutaminyl-
l
-lysine endopeptidase activity. The endopeptidase cleavage specificity of the lysins was confirmed using a synthetic peptide substrate corresponding to a portion of the stem peptide and cross bridge of
Streptococcus agalactiae
peptidoglycan. The LambdaSa2 lysin also displayed β-
d
-
N
-acetylglucosaminidase activity.
Activation of Endocytosis as an Adaptation to the Mammalian Host by Trypanosomes
Authors: Natesan Senthil Kumar A. , Peacock Lori , Matthews Keith , Gibson Wendy , Field Mark C. 
Published: September 2007

ABSTRACT

Immune evasion in African trypanosomes is principally mediated by antigenic variation, but rapid internalization of surface-bound immune factors may contribute to survival. Endocytosis is upregulated approximately 10-fold in bloodstream compared to procyclic forms, and surface coat remodeling accompanies transition between these life stages. Here we examined expression of endocytosis markers in tsetse fly stages in vivo and monitored modulation during transition from bloodstream to procyclic forms in vitro. Among bloodstream stages nonproliferative stumpy forms have endocytic activity similar to that seen with rapidly dividing slender forms, while differentiation of stumpy forms to procyclic forms is accompanied by rapid down-regulation of Rab11 and clathrin, suggesting that modulation of endocytic and recycling systems accompanies this differentiation event. Significantly, rapid down-regulation of endocytic markers occurs upon entering the insect midgut and expression of Rab11 and clathrin remains low throughout subsequent development, which suggests that high endocytic activity is not required for remodeling the parasite surface or for survival within the fly. However, salivary gland metacyclic forms dramatically increase expression of clathrin and Rab11, indicating that emergence of mammalian infective forms is coupled to reacquisition of a high-activity endocytic-recycling system. These data suggest that high-level endocytosis in
Trypanosoma brucei
is an adaptation required for viability in the mammalian host.
O
-Mannosyltransferase 1 in
Aspergillus fumigatus
(AfPmt1p) Is Crucial for Cell Wall Integrity and Conidium Morphology, Especially at an Elevated Temperature
Authors: Zhou Hui , Hu Hongyan , Zhang Lijuan , Li Ruoyu , Ouyang Haomiao , Ming Jia , Jin Cheng 
Published: September 2007

ABSTRACT

Protein O-mannosyltransferases initiate O mannosylation of secretory proteins, which are of fundamental importance in eukaryotes. In this study, the
PMT
gene family of the human fungal pathogen
Aspergillus fumigatus
was identified and characterized. Unlike the case in
Saccharomyces cerevisiae
, where the
PMT
family is highly redundant, only one member of each
PMT
subfamily, namely, Af
pmt1
, Af
pmt2
, and Af
pmt4
, is present in
A. fumigatus
. Mutants with a deletion of Af
pmt1
are viable. In vitro and in vivo activity assays confirmed that the protein encoded by Af
pmt1
acts as an O-mannosyltransferase (AfPmt1p). Characterization of the ΔAf
pmt1
mutant showed that a lack of AfPmt1p results in sensitivity to elevated temperature and defects in growth and cell wall integrity, thereby affecting cell morphology, conidium formation, and germination. In a mouse model, Af
pmt1
was not required for the virulence of
A. fumigatus
under the experimental conditions used.
Functional Characterization of the Fission Yeast Phosphatidylserine Synthase Gene,
pps1
, Reveals Novel Cellular Functions for Phosphatidylserine
Authors: Matsuo Yasuhiro , Fisher Edward , Patton-Vogt Jana , Marcus Stevan 
Published: September 2007

ABSTRACT

To investigate the contributions of phosphatidylserine to the growth and morphogenesis of the rod-shaped fission yeast
Schizosaccharomyces pombe
, we have characterized the single gene in this organism,
pps1
, encoding a predicted phosphatidylserine synthase.
S. pombe pps1
Δ mutants grow slowly in rich medium and are inviable in synthetic minimal medium. They do not produce detectable phosphatidylserine in vivo and possess negligible in vitro phosphatidylserine synthase activity, indicating that
pps1
encodes the major phosphatidylserine synthase activity in
S. pombe
. Supplementation of growth medium with ethanolamine partially suppresses the growth-defective phenotype of
pps1
Δ cells, reflecting the likely importance of phosphatidylserine as a precursor for phosphatidylethanolamine in
S. pombe
. In medium lacking ethanolamine,
pps1
Δ mutants exhibit striking cell morphology, cytokinesis, actin cytoskeleton, and cell wall remodeling and integrity defects. Overexpression of
pps1
likewise leads to defects in cell morphology and cytokinesis, thus implicating phosphatidylserine as a dosage-dependent regulator of these processes. During log-phase growth, green fluorescent protein-Pps1p fusion proteins are concentrated at the cell and nuclear peripheries as well as presumptive endoplasmic reticulum membranes, while in stationary-phase cells, they are redistributed to unusual cytoplasmic structures of unknown origin. Moreover, stationary-phase
pps1
Δ cultures retain very poor viability relative to wild-type
S. pombe
cells, even in medium containing ethanolamine, demonstrating a role for phosphatidylserine in the physiological adaptations required for stationary-phase survival. Our findings reveal novel cellular functions for phosphatidylserine and emphasize the usefulness of
S. pombe
as a model organism for elucidating potentially conserved biological and molecular functions of this phospholipid.
Sexual Pheromone Induces Diffusion of the Pheromone-Homologous Polypeptide in the Extracellular Matrix of
Volvox carteri
Authors: Ishida Koichi 
Published: September 2007

ABSTRACT

The C-terminal domain of pherophorin II is homologous to the sexual pheromone of
Volvox carteri
and is released from other domains during sexual induction. Green fluorescent protein fused to the C terminus of pherophorin II was located at the extracellular matrix directly surrounding the gonidium, the final target of the sexual-induction signal.
Two Iron-Responsive Promoter Elements Control Expression of
FOX1
in
Chlamydomonas reinhardtii
Authors: Deng Xiaodong , Eriksson Mats 
Published: September 2007

ABSTRACT

FOX1
encodes an iron deficiency-induced ferroxidase involved in a high-affinity iron uptake system. Mutagenesis analysis of the
FOX1
promoter identified two separate iron-responsive elements, FeRE1 (CACACG) and FeRE2 (CACGCG), between positions −87 and −82 and between positions −65 and −60, respectively, and both are needed for induced
FOX1
expression under conditions of iron deficiency.
Biochemical Characterization of dTDP-
d
-Qui4N and dTDP-
d
-Qui4NAc Biosynthetic Pathways in
Shigella dysenteriae
Type 7 and
Escherichia coli
O7
Authors: Wang Ying , Xu Yanli , Perepelov Andrei V. , Qi Yuanyuan , Knirel Yuriy A. , Wang Lei , Feng Lu 
Published: September 2007

ABSTRACT

O-antigen variation due to the presence of different types of sugars and sugar linkages is important for the survival of bacteria threatened by host immune systems. The O antigens of
Shigella dysenteriae
type 7 and
Escherichia coli
O7 contain 4-(
N
-acetylglycyl)amino-4,6-dideoxy-
d
-glucose (
d
-Qui4NGlyAc) and 4-acetamido-4,6-dideoxy-
d
-glucose (
d
-Qui4NAc), respectively, which are sugars not often found in studied polysaccharides. In this study, we characterized the biosynthetic pathways for dTDP-
d
-Qui4N and dTDP-
d
-Qui4NAc (the nucleotide-activated precursors of
d
-Qui4NGlyAc and
d
-Qui4NAc in O antigens). Predicted genes involved in the synthesis of the two sugars were cloned, and the gene products were overexpressed and purified as His-tagged fusion proteins. In vitro enzymatic reactions were carried out using the purified proteins, and the reaction products were analyzed by capillary electrophoresis, electrospray ionization-mass spectrometry, and nuclear magnetic resonance spectroscopy. It is shown that in
S. dysenteriae
type 7 and
E. coli
O7, dTDP-
d
-Qui4N is synthesized from α-
d
-glucose-1-phosphate in three reaction steps catalyzed by glucose-1-phosphate thymidyltransferase (RmlA), dTDP-
d
-glucose 4,6-dehydratase (RmlB), and dTDP-4-keto-6-deoxy-
d
-glucose aminotransferase (VioA). An additional acetyltransferase (VioB) catalyzes the conversion of dTDP-
d
-Qui4N into dTDP-
d
-Qui4NAc in
E. coli
O7. Kinetic parameters and some other properties of VioA and VioB are described and differences between VioA proteins from
S. dysenteriae
type 7 (VioA
D7
) and
E. coli
O7 (VioA
O7
) discussed. To our knowledge, this is the first time that functions of VioA and VioB have been biochemically characterized. This study provides valuable enzyme sources for the production of dTDP-
d
-Qui4N and dTDP-
d
-Qui4NAc, which are potentially useful in the pharmaceutical industry for drug development.
Sequencing the Botulinum Neurotoxin Gene and Related Genes in
Clostridium botulinum
Type E Strains Reveals
orfx3
and a Novel Type E Neurotoxin Subtype
Authors: Chen Ying , Korkeala Hannu , Aarnikunnas Johannes , Lindström Miia 
Published: September 2007

ABSTRACT

Three
Clostridium botulinum
type E strains were sequenced for the botulinum neurotoxin (BoNT) gene cluster, and 11 type E strains, representing a wide biodiversity, were sequenced for the
bont
/E gene. The total length of the BoNT/E gene cluster was 12,908 bp, and a novel gene (partial) designated
orfx3
, together with the complete
orfx2
gene, was identified in the three type E strains for the first time. Apart from
orfx3
, the structure and organization of the neurotoxin gene cluster of the three strains were identical to those of previously published ones. Only minor differences (≤3%) in the nucleotide sequences of the gene cluster components were observed among the three strains and the published BoNT/E-producing clostridia. The
orfx3
,
orfx2
,
orfx1
, and
p47
gene sequences of the three type E strains shared homologies of 81%, 67 to 76%, 78 to 79%, and 79 to 85%, respectively, with published sequences for type A1 and A2
C. botulinum
. Analysis of
bont
/E from the 14 type E strains and 19 previously published BoNT/E-producing clostridia revealed six neurotoxin subtypes, with a new distinct subtype consisting of three Finnish isolates alone. The amino acid sequence of the subtype E6 neurotoxin differed 3 to 6% from the other subtypes, suggesting that these subtype E6 neurotoxins may possess specific antigenic or functional properties.
In Vivo Mutations of Thymidylate Synthase (Encoded by
thyA
) Are Responsible for Thymidine Dependency in Clinical Small-Colony Variants of
Staphylococcus aureus
Authors: Chatterjee Indranil , Kriegeskorte Andre , Fischer Andreas , Deiwick Susanne , Theimann Nadine , Proctor Richard A. , Peters Georg , Herrmann Mathias , Kahl Barbara C. 
Published: September 2007

ABSTRACT

Trimethoprim-sulfamethoxazole (SXT)-resistant
Staphylococcus aureus
thymidine-dependent small-colony variants (TD-SCVs) are frequently isolated from the airways of cystic fibrosis (CF) patients, often in combination with isogenic normal strains if patients were treated with SXT for extended periods. As SXT inhibits the synthesis of tetrahydrofolic acid, which acts as a cofactor for thymidylate synthase (
thyA
), the survival of TD-SCVs depends exclusively on the availability of external thymidine. Since the underlying mechanism for thymidine dependency is unknown, we investigated if alterations in the
thyA
nucleotide sequences were responsible for this phenomenon. Sequence analysis of several clinical TD-SCVs and their isogenic normal strains with reference to previously published
S. aureus thyA
nucleotide sequences was performed. Three clinical TD-SCVs were complemented by transforming TD-SCVs with the vector pCX19 expressing ThyA from
S. aureus
8325-4. Transcriptional analysis of metabolic and virulence genes and regulators (
agr
,
hla
,
spa
,
citB
,
thyA
, and
nupC
) was performed by quantitative reverse transcription-PCR. The previously published sequences of
thyA
and two normal clinical strains were highly conserved, while
thyA
of four normal strains and four SCVs had nonsynonymous point mutations. In 8/10 SCVs, deletions occurred, resulting in stop codons which were located in 4/10 SCVs close to or within the active site of the protein (dUMP binding). Complementation of TD-SCVs with
thyA
almost fully reversed the phenotype, growth characteristics, and transcription patterns. In conclusion, we demonstrated that mutations of the
thyA
gene were responsible for the phenotype of TD-SCVs. Complementation of TD-SCVs with
thyA
revealed that a functional ThyA protein is necessary and sufficient to change the SCV phenotype and behavior back to normal.
A New O-Antigen Gene Cluster Has a Key Role in the Virulence of the
Escherichia coli
Meningitis Clone O45:K1:H7
Authors: Plainvert Céline , Bidet Philippe , Peigne Chantal , Barbe Valérie , Médigue Claudine , Denamur Erick , Bingen Edouard , Bonacorsi Stéphane 
Published: September 2007

ABSTRACT

A new highly pathogenic clone of
Escherichia coli
meningitis strains harboring the unusual serogroup O45 has recently emerged in France. To gain insight into the pathogenicity of this new clone, we investigated the possible role of antigen O45 in the virulence of strain S88 (O45:K1:H7), representative of this emerging clone. We first showed that the S88 O-antigen gene cluster sequence differs from that of O45 in the reference strain
E. coli
96-3285, suggesting that the two O45 polysaccharides, while probably sharing a community of epitopes, represent two different antigens. The unique functional organization of the two O-antigen gene clusters and the low DNA sequence homology of the orthologous genes suggest that the two loci originated from a common ancestor and have since undergone multiple recombination events. Phylogenetic analysis based on the flanking gene
gnd
sequences indicates that the S88 antigen O45 (O45
S88
) gene cluster may have been acquired, at least in part, from another member of the
Enterobacteriaceae
. Mutagenesis of the O45
S88
antigen gene cluster was used for functional analysis of the loci and revealed the crucial role of the O polysaccharide in S88 virulence in a neonatal rat meningitis model. We also developed a PCR method to specifically identify the O45
S88
antigen gene cluster. Together, our findings suggest that horizontal acquisition of a new O-antigen gene cluster, at least partly from another species, may have been a key event in the emergence and virulence of the
E. coli
O45:K1:H7 clone in France.
Response of the Hyperthermophilic ArchaeonSulfolobus solfataricusto UV Damage
Authors: Fröls Sabrina , Gordon Paul M. K. , Panlilio Mayi Arcellana , Duggin Iain G. , Bell Stephen D. , Sensen Christoph W. , Schleper Christa 
Published: September 2007

ABSTRACTIn order to characterize the genome-wide transcriptional response of the hyperthermophilic, aerobic crenarchaeoteSulfolobus solfataricusto UV damage, we used high-density DNA microarrays which covered 3,368 genetic features encoded on the host genome, as well as the genes of several extrachromosomal genetic elements. While no significant up-regulation of genes potentially involved in direct DNA damage reversal was observed, a specific transcriptional UV response involving 55 genes could be dissected. Although flow cytometry showed only modest perturbation of the cell cycle, strong modulation of the transcript levels of the Cdc6 replication initiator genes was observed. Up-regulation of an operon encoding Mre11 and Rad50 homologs pointed to induction of recombinational repair. Consistent with this, DNA double-strand breaks were observed between 2 and 8 h after UV treatment, possibly resulting from replication fork collapse at damaged DNA sites. The strong transcriptional induction of genes which potentially encode functions for pilus formation suggested that conjugational activity might lead to enhanced exchange of genetic material. In support of this, a statistical microscopic analysis demonstrated that large cell aggregates formed upon UV exposure. Together, this provided supporting evidence to a link between recombinational repair and conjugation events.