bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of Protein-Protein Interactions Involved in Iron Reduction by
Shewanella oneidensis
MR-1
Authors: Ross Daniel E. , Ruebush Shane S. , Brantley Susan L. , Hartshorne Robert S. , Clarke Thomas A. , Richardson David J. , Tien Ming 
Published: August 2007

ABSTRACT

The interaction of proteins implicated in dissimilatory metal reduction by
Shewanella oneidensis
MR-1 (outer membrane [OM] proteins OmcA, MtrB, and MtrC; OM-associated protein MtrA; periplasmic protein CctA; and cytoplasmic membrane protein CymA) were characterized by protein purification, analytical ultracentrifugation, and cross-linking methods. Five of these proteins are heme proteins, OmcA (83 kDa), MtrC (75 kDa), MtrA (32 kDa), CctA (19 kDa), and CymA (21 kDa), and can be visualized after sodium dodecyl sulfate-polyacrylamide gel electrophoresis by heme staining. We show for the first time that MtrC, MtrA, and MtrB form a 198-kDa complex with a 1:1:1 stoichiometry. These proteins copurify through anion-exchange chromatography, and the purified complex has the ability to reduce multiple forms of Fe(III) and Mn(IV). Additionally, MtrA fractionates with the OM through sucrose density gradient ultracentrifugation, and MtrA comigrates with MtrB in native gels. Protein cross-linking of whole cells with 1% formaldehyde show new heme bands of 160, 151, 136, and 59 kDa. Using antibodies to detect each protein separately, heme proteins OmcA and MtrC were shown to cross-link, yielding the 160-kDa band. Consistent with copurification results, MtrB cross-links with MtrA, forming high-molecular-mass bands of approximately 151 and 136 kDa.
Outcome of Colonization of
Apis mellifera
by
Nosema ceranae
Authors: Martín-Hernández Raquel , Meana Aránzazu , Prieto Lourdes , Salvador Amparo Martínez , Garrido-Bailón Encarna , Higes Mariano 
Published: August 2007

ABSTRACT

A multiplex PCR-based method, in which two small-subunit rRNA regions are simultaneously amplified in a single reaction, was designed for parallel detection of honeybee microsporidians (
Nosema apis
and
Nosema ceranae
). Each of two pairs of primers exclusively amplified the 16S rRNA targeted gene of a specific microsporidian. The multiplex PCR assay was useful for specific detection of the two species of microsporidians related to bee nosemosis, not only in purified spores but also in honeybee homogenates and in naturally infected bees. The multiplex PCR assay was also able to detect coinfections by the two species. Screening of bee samples from Spain, Switzerland, France, and Germany using the PCR technique revealed a greater presence of
N. ceranae
than of
N. apis
in Europe, although both species are widely distributed. From the year 2000 onward, statistically significant differences have been found in the proportions of
Nosema
spp. spore-positive samples collected between and within years. In the first period examined (1999 to 2002), the smallest number of samples diagnosed as
Nosema
positive was found during the summer months, showing clear seasonality in the diagnosis, which is characteristic of
N. apis
. From 2003 onward a change in the tendency resulted in an increase in
Nosema
-positive samples in all months until 2005, when a total absence of seasonality was detected. A significant causative association between the presence of
N. ceranae
and hive depopulation clearly indicates that the colonization of
Apis mellifera
by
N. ceranae
is related to bee losses.
Use of Cell Culture To Assess
Cryptosporidium parvum
Survival Rates in Natural Groundwaters and Surface Waters
Authors: Ives Rebecca L. , Kamarainen Amy M. , John David E. , Rose Joan B. 
Published: August 2007

ABSTRACT

Bench-scale survival studies with
Cryptosporidium parvum
were conducted with representative aquifer and reservoir waters of Florida.
C. parvum
inactivation rates ranged from 0.0088 log
10
/day at 5°C to −0.20 log
10
/day at 30°C. Temperature, surface water or groundwater type, and the interaction of these factors had statistically significant effects on the survival of
C. parvum
.
agr
System of
Listeria monocytogenes
EGD-e: Role in Adherence and Differential Expression Pattern
Authors: Rieu Aurélie , Weidmann Stéphanie , Garmyn Dominique , Piveteau Pascal , Guzzo Jean 
Published: August 2007

ABSTRACT

In this study, we investigated the
agrBDCA
operon in the pathogenic bacterium
Listeria monocytogenes
EGD-e. In-frame deletion of
agrA
and
agrD
resulted in an altered adherence and biofilm formation on abiotic surfaces, suggesting the involvement of the
agr
system of
L. monocytogenes
during the early stages of biofilm formation. Real-time PCR experiments indicated that the transcript levels of
agrBDCA
depended on the stage of biofilm development, since the levels were lower after the initial attachment period than during biofilm growth, whereas transcription during planktonic growth was not growth phase dependent. The mRNA quantification data also suggested that the
agr
system was autoregulated and pointed to a differential expression of the
agr
genes during sessile and planktonic growth. Although the reverse transcription-PCR experiments revealed that the four genes were transcribed as a single messenger, chemical half-life and 5′ RACE (rapid amplification of cDNA ends) experiments indicated that the full size transcript underwent cleavage followed by degradation of the
agrC
and
agrA
transcripts, which suggests a complex regulation of
agr
transcription.
Cloning and Transfer of the
Salmonella
Pathogenicity Island 2 Type III Secretion System for Studies of a Range of Gram-Negative Genera
Authors: Wilson James W. , Coleman Clint , Nickerson Cheryl A. 
Published: August 2007

ABSTRACT

The engineering of bacterial strains with specific phenotypes frequently requires the use of blocks or “cassettes” of genes that act together to perform a desired function. The potential benefits of utilizing type III secretion systems in this regard are becoming increasingly realized since these systems can be used to direct interactions with host cells for beneficial purposes such as vaccine development, anticancer therapies, and targeted protein delivery. However, convenient methods to clone and transfer type III secretion systems for studies of a range of different types of bacteria are lacking. In addition to functional applications, such methods would also reveal important information about the evolution of a given type III secretion system, such as its ability to be expressed and functional outside of the strain of origin. We describe here the cloning of the
Salmonella enterica
serovar Typhimurium pathogenicity island 2 (SPI-2) type III secretion system onto a vector that can be easily transferred to a range of gram-negative bacterial genera. We found that expression of the cloned SPI-2 system in different
Gammaproteobacteria
and
Alphaproteobacteria
(as monitored by SseB protein levels) is dependent on the bacterial strain and growth medium. We also demonstrate that the cloned system is functional for secretion, can direct interactions with macrophages, and can be used as a novel tool to analyze the predicted interaction of SseB with host cells. This work provides a foundation for future applications where the cloned SPI-2 region (or other cloned type III systems) can provide a desired function to an engineered gram-negative strain.
Molecular Characterization of the
phaEC
Hm
Genes, Required for Biosynthesis of Poly(3-Hydroxybutyrate) in the Extremely Halophilic Archaeon
Haloarcula marismortui
Authors: Han Jing , Lu Qiuhe , Zhou Ligang , Zhou Jian , Xiang Hua 
Published: August 2007

ABSTRACT

Although many haloarchaea produce biodegradable polyhydroxyalkanoates (PHAs), the genes involved in PHA synthesis in the domain of
Archaea
have not yet been experimentally investigated yet. In this study, we revealed that
Haloarcula marismortui
was able to accumulate poly(3-hydroxybutyrate) (PHB) up to 21% of cellular dry weight when cultured in a minimal medium with excessive glucose and identified the
phaE
Hm
and
phaC
Hm
genes, probably encoding two subunits of a class III PHA synthase. These two genes were adjacent and directed by a single promoter located 26 bp upstream of the transcriptional start site and were constitutively expressed under both nutrient-rich and -limited conditions. Interestingly, PhaC
Hm
was revealed to be strongly bound with the PHB granules, but PhaE
Hm
seemed not to be. Introduction of either the
phaE
Hm
or
phaC
Hm
gene into
Haloarcula hispanica
, which harbors highly homologous
phaEC
Hh
genes, could enhance the PHB synthesis in the recombinant strains, while coexpression of the both genes always generated the highest PHB yield. Significantly, knockout of the
phaEC
Hh
genes in
H. hispanica
led to a complete loss of the PHA synthase activity. Complementation with
phaEC
Hm
genes, but not a single one, restored the capability of PHB accumulation as well as the PHA synthase activity in this
phaEC
-deleted haloarchaeon. These results indicated that the
phaEC
genes are required for biosynthesis of PHB and might encode an active PHA synthase in the
Haloarcula
species.
Transcriptome Analysis ofPseudomonas putidaKT2440 Harboring the Completely Sequenced IncP-7 Plasmid pCAR1
Authors: Miyakoshi Masatoshi , Shintani Masaki , Terabayashi Tsuguno , Kai Satoshi , Yamane Hisakazu , Nojiri Hideaki 
Published: August 2007

ABSTRACTThe IncP-7 plasmid pCAR1 ofPseudomonas resinovoransCA10 confers the ability to degrade carbazole upon transfer to the recipient strainP. putidaKT2440. We designed a customized whole-genome oligonucleotide microarray to study the coordinated expression of pCAR1 and the chromosome in the transconjugant strain KT2440(pCAR1). First, the transcriptome of KT2440(pCAR1) during growth with carbazole as the sole carbon source was compared to that during growth with succinate. The carbazole cataboliccarandantoperons were induced, along with the chromosomalcatandpcagenes involved in the catechol branch of the β-ketoadipate pathway. Additionally, the regulatory geneantRencoding the AraC/XylS family transcriptional activator specific forcarandantoperons was upregulated. The characterization of theantRpromoter revealed thatantRis transcribed from an RpoN-dependent promoter, suggesting that the successful expression of the carbazole catabolic operons depends on whether the chromosome contains the specific RpoN-dependent activator. Next, to analyze whether the horizontal transfer of a plasmid alters the transcription network of its host chromosome, we compared the chromosomal transcriptomes of KT2440(pCAR1) and KT2440 under the same growth conditions. Only subtle changes were caused by the transfer of pCAR1, except for the significant induction of the hypothetical gene PP3700, designatedparI, which encodes a putative ParA-like ATPase with an N-terminal Xre-type DNA-binding motif. Further transcriptional analyses showed that theparIpromoter was positively regulated by ParI itself and the pCAR1-encoded protein ParA.
Comparative Genomic Analyses of Seventeen
Streptococcus pneumoniae
Strains: Insights into the Pneumococcal Supragenome
Authors: Hiller N. Luisa , Janto Benjamin , Hogg Justin S. , Boissy Robert , Yu Susan , Powell Evan , Keefe Randy , Ehrlich Nathan E. , Shen Kai , Hayes Jay , Barbadora Karen , Klimke William , Dernovoy Dmitry , Tatusova Tatiana , Parkhill Julian , Bentley Stephen D. , Post J. Christopher , Ehrlich Garth D. , Hu Fen Z. 
Published: August 2007

ABSTRACT

The distributed-genome hypothesis (DGH) states that pathogenic bacteria possess a supragenome that is much larger than the genome of any single bacterium and that these pathogens utilize genetic recombination and a large, noncore set of genes as a means of diversity generation. We sequenced the genomes of eight nasopharyngeal strains of S
treptococcus pneumoniae
isolated from pediatric patients with upper respiratory symptoms and performed quantitative genomic analyses among these and nine publicly available pneumococcal strains. Coding sequences from all strains were grouped into 3,170 orthologous gene clusters, of which 1,454 (46%) were conserved among all 17 strains. The majority of the gene clusters, 1,716 (54%), were not found in all strains. Genic differences per strain pair ranged from 35 to 629 orthologous clusters, with each strain's genome containing between 21 and 32% noncore genes. The distribution of the orthologous clusters per genome for the 17 strains was entered into the finite-supragenome model, which predicted that (i) the
S. pneumoniae
supragenome contains more than 5,000 orthologous clusters and (ii) 99% of the orthologous clusters (∼3,000) that are represented in the
S. pneumoniae
population at frequencies of ≥0.1 can be identified if 33 representative genomes are sequenced. These extensive genic diversity data support the DGH and provide a basis for understanding the great differences in clinical phenotype associated with various pneumococcal strains. When these findings are taken together with previous studies that demonstrated the presence of a supragenome for
Streptococcus agalactiae
and
Haemophilus influenzae
, it appears that the possession of a distributed genome is a common host interaction strategy.
The Hotdog Thioesterase EntH (YbdB) Plays a Role In Vivo in Optimal Enterobactin Biosynthesis by Interacting with the ArCP Domain of EntB
Authors: Leduc Damien , Battesti Aurélia , Bouveret Emmanuelle 
Published: August 2007

ABSTRACT

In response to iron limitation, the siderophore enterobactin is synthesized and secreted by
Escherichia coli
. Its biosynthesis is performed by a series of enzymes encoded by the Ent gene cluster. Among the genes of this cluster,
ybdB
has not been implicated in enterobactin production to date. We demonstrate here an in vivo role for the hotdog protein EntH (YbdB) in the optimal production of enterobactin. Indeed, we showed that EntH is a thioesterase specifically produced under iron limitation conditions. Furthermore, EntH interacts specifically with the aryl carrier protein (ArCP) domain of EntB, a crucial bifunctional enzyme of the enterobactin biosynthesis pathway and a potential target of EntH thioesterase activity. A strain devoid of EntH is impaired for growth under iron limitation associated with the presence of the salicylate inhibitor, correlating with the diminution of enterobactin production under these conditions. Normal growth and enterobactin production are restored upon expression of
entH
in
trans
. Inversely, unnecessary overproduction of EntH provokes a fall of the quantity of siderophore produced under iron starvation conditions. Our findings point to a proofreading role for EntH during biosynthesis of enterobactin in vivo. EntH thioesterase activity could be required for cleaving wrongly charged molecules on the carrier protein EntB. This is the first description of such a role in the optimization of a nonribosomal biosynthesis pathway for a protein of the hotdog superfamily.
Generation of Virulence Factor Variants in
Staphylococcus aureus
Biofilms
Authors: Yarwood Jeremy M. , Paquette Kara M. , Tikh Ilya B. , Volper Esther M. , Greenberg E. Peter 
Published: August 2007

ABSTRACT

Several serious diseases are caused by biofilm-associated
Staphylococcus aureus
. Colonial variants occur in biofilms of other bacterial species, and
S. aureus
variants are frequently isolated from biofilm-associated infections. Thus, we studied the generation of variants with altered expression of virulence factors in
S. aureus
biofilms. We observed that the number of variants found in biofilms, as measured by hemolytic activity, varied for different strains. Further study of hemolytic activity and signaling by the accessory gene regulator (Agr) quorum-sensing system in one
S. aureus
strain revealed three primary biofilm subpopulations: nonhemolytic (Agr deficient), hemolytic (Agr positive), and hyperhemolytic (also Agr positive). The nonhemolytic variant became the numerically dominant subpopulation in the biofilm. The nonhemolytic variant phenotype was stable and heritable, indicating a genetic perturbation, whereas the hyperhemolytic phenotype was unstable, suggesting a phase variation. Transcription profiling revealed that expression of the
agr
locus and many extracellular virulence factors was repressed in the nonhemolytic variant. Expression of the
agr
-activating gene,
sarU
, was also repressed in the nonhemolytic variant, suggesting one potential regulatory pathway responsible for the Agr-deficient phenotype. We suggest that the development of these variants in biofilms may have important clinical implications.
Transcriptomic Analysis of the Sulfate Starvation Response ofPseudomonas aeruginosa
Authors: Tralau Tewes , Vuilleumier Stéphane , Thibault Christelle , Campbell Barry J. , Hart C. Anthony , Kertesz Michael A. 
Published: August 2007

ABSTRACTPseudomonas aeruginosais an opportunistic pathogen that causes a number of infections in humans, but is best known for its association with cystic fibrosis. It is able to use a wide range of sulfur compounds as sources of sulfur for growth. Gene expression in response to changes in sulfur supply was studied inP. aeruginosaE601, a cystic fibrosis isolate that displays mucin sulfatase activity, and inP. aeruginosaPAO1. A large family of genes was found to be upregulated by sulfate limitation in both isolates, encoding sulfatases and sulfonatases, transport systems, oxidative stress proteins, and a sulfate-regulated TonB/ExbBD complex. These genes were localized in five distinct islands on the genome and encoded proteins with a significantly reduced content of cysteine and methionine. Growth ofP. aeruginosaE601 with mucin as the sulfur source led not only to a sulfate starvation response but also to induction of genes involved with type III secretion systems.
Regulatory Overlap and Functional Redundancy amongBacillus subtilisExtracytoplasmic Function σ Factors
Authors: Mascher Thorsten , Hachmann Anna-Barbara , Helmann John D. 
Published: August 2007

ABSTRACTBacillus subtilis encodes seven extracytoplasmic function (ECF) σ factors that regulate partially overlapping regulons related to cell envelope homeostasis and antibiotic resistance. Here, we investigated their physiological role by constructing a mutant set of single, double, triple, and quadruple ECF σ factor deletions in the undomesticatedB. subtilisstrain NCIB3610. This mutant set was subsequently screened for defects in motility, multicellular differentiation, and sensitivity to more than 200 chemicals by using Phenotype MicroArrays. A quadruple mutant strain, harboring deletions of thesigV,sigY,sigZ, andylaCgene, behaved indistinguishably from the wild-type strain, indicative of either regulatory redundancy or very specific functions of these four ECF σ factors. In contrast, a triple mutant, inactivated for thesigM,sigW, andsigXgenes (but none of the corresponding double mutants), showed a biphasic growth behavior and a complete loss of multicellular differentiation, as judged by both colony formation and the inability to form a pellicle. This triple mutant also displayed a greatly increased sensitivity to detergents and several cell wall antibiotics including β-lactams, polymyxin B, andd-cycloserine. In several cases, these antibiotic-sensitive phenotypes are significantly enhanced in the triple mutant strain relative to strains lacking only one or two σ factors.
Role of Nucleoid-Associated Proteins Hha and H-NS in Expression of
Salmonella enterica
Activators HilD, HilC, and RtsA Required for Cell Invasion
Authors: Olekhnovich Igor N. , Kadner Robert J. 
Published: August 2007

ABSTRACT

The coordinate expression of
Salmonella enterica
invasion genes on
Salmonella
pathogenicity island 1 is under the control of the complex circuits of regulation that involve the AraC/XylS family transcriptional activators HilD, HilC, and RtsA and nucleoid-associated proteins. Single-copy transcription fusions were used to assess the effects of nucleoid-associated proteins Hha and H-NS on
hilD
,
hilC
, and
rtsA
expression. The data show that all three genes,
hilD
,
hilC
, and
rtsA
, were repressed by H-NS and/or Hha. The repression of
rtsA
was the highest among tested genes. The level of
rtsA-lac
was equally elevated in
hns
and
hha
mutants and was further enhanced in the
hns hha
double mutant under low-osmolarity conditions. Electrophoretic mobility shift experiments showed that H-NS and Hha directly bind to the
rtsA
promoter. In addition to the negative control that was exerted by H-NS/Hha under low-osmolarity conditions, the homologous virulence activators HilD, HilC, and RtsA (Hil activators) induced
rtsA-lac
expression in a high-salt medium. A DNase footprinting assay of the
rtsA
promoter revealed one common DNA-binding site for all three Hil activators centered at position −54 relative to the transcriptional start site. In the absence of Hha and H-NS, however, osmoregulation of the
rtsA
promoter was lost, and Hil activators were not required for
rtsA
transcription. These results taken together suggest that the HilD, HilC, and RtsA proteins induce the transcription of the
rtsA
promoter by counteracting H-NS/Hha-mediated repression.
High-risk human papillomavirus is present in cytologically false-negative smears: an analysis of “normal” smears preceding CIN2/3
Authors: Bulk S , Rozendaal L , Zielinski G D , Berkhof J , Daalmeijer N C Fransen , Snijders P J F , van Kemenade F J , Meijer C J L M 
Published: August 2007

Aims:Cervical screening, currently performed by cervical cytology, depends on the timely detection of malignant lesions for its success. The presence of high-risk human papillomavirus (hrHPV) is associated with an increased risk of subsequent high-grade cervical intra-epithelial neoplasia (CIN2/3) and cervical cancer. The aim of this study was to determine the extent to which hrHPV is present in cervical smears with a high a priori chance of being false negative (ie, in normal smears preceding CIN2/3).Methods:Archival specimens of 187 women with CIN2/3 and preceding normal conventional smears were identified retrospectively. Of these specimens, 144 (77%) had adequate cytological samples for further HPV DNA testing.Results:Of 144 CIN2/3 lesions, preceding normal smears showed hrHPV positivity in 80% of cases. Of the hrHPV-positive smears, 69% were upgraded cytologically at rescreening compared with 24% of hrHPV-negative smears (p<0.001). Upgrading of smears was not associated with specific hrHPV types (p = 0.217). In over 90% of cases, type concordance in smear and CIN2/3 lesion was demonstrated.Conclusions:hrHPV is present in a high proportion of normal archival smears preceding CIN2/3, and false-negative cytology was highly associated with the presence of hrHPV. This supports the current notion that hrHPV testing can be used as a primary cervical screening tool. If so, hrHPV-positive cervical smears should be carefully examined for cytological abnormalities to reduce false-negative cervical cytology.