bluuebunny/crossref_metadata_2025 huggingface dataset:
Novel Pyrophosphate-Forming Acetate Kinase from the Protist Entamoeba histolytica
Authors: Fowler Matthew L. , Ingram-Smith Cheryl , Smith Kerry S.
Published: August 2012

Acetate kinase (ACK) catalyzes the reversible synthesis of acetyl phosphate by transfer of the γ-phosphate of ATP to acetate. Here we report the first biochemical and kinetic characterization of a eukaryotic ACK, that from the protist
Entamoeba histolytica
. Our characterization revealed that this protist ACK is the only known member of the ASKHA structural superfamily, which includes acetate kinase, hexokinase, and other sugar kinases, to utilize inorganic pyrophosphate (PP
i
)/inorganic phosphate (P
i
) as the sole phosphoryl donor/acceptor. Detection of ACK activity in
E. histolytica
cell extracts in the direction of acetate/PP
i
formation but not in the direction of acetyl phosphate/P
i
formation suggests that the physiological direction of the reaction is toward acetate/PP
i
production. Kinetic parameters determined for each direction of the reaction are consistent with this observation. The
E. histolytica
PP
i
-forming ACK follows a sequential mechanism, supporting a direct in-line phosphoryl transfer mechanism as previously reported for the well-characterized
Methanosarcina thermophila
ATP-dependent ACK. Characterizations of enzyme variants altered in the putative acetate/acetyl phosphate binding pocket suggested that acetyl phosphate binding is not mediated solely through a hydrophobic interaction but also through the phosphoryl group, as for the
M. thermophila
ACK. However, there are key differences in the roles of certain active site residues between the two enzymes. The absence of known ACK partner enzymes raises the possibility that ACK is part of a novel pathway in
Entamoeba
.
A Distinct Regulatory Sequence Is Essential for the Expression of a Subset of
nle
Genes in Attaching and Effacing Escherichia coli
Authors: García-Angulo Víctor A. , Martínez-Santos Verónica I. , Villaseñor Tomás , Santana Francisco J. , Huerta-Saquero Alejandro , Martínez Luary C. , Jiménez Rafael , Lara-Ochoa Cristina , Téllez-Sosa Juan , Bustamante Víctor H. , Puente José L.
Published: August 2012

Enteropathogenic
Escherichia coli
uses a type III secretion system (T3SS), encoded in the locus of enterocyte effacement (LEE) pathogenicity island, to translocate a wide repertoire of effector proteins into the host cell in order to subvert cell signaling cascades and promote bacterial colonization and survival. Genes encoding type III-secreted effectors are located in the LEE and scattered throughout the chromosome. While LEE gene regulation is better understood, the conditions and factors involved in the expression of effectors encoded outside the LEE are just starting to be elucidated. Here, we identified a highly conserved sequence containing a 13-bp inverted repeat (IR), located upstream of a subset of genes coding for different non-LEE-encoded effectors in A/E pathogens. Site-directed mutagenesis and deletion analysis of the
nleH1
and
nleB2
regulatory regions revealed that this IR is essential for the transcriptional activation of both genes. Growth conditions that favor the expression of LEE genes also facilitate the activation of
nleH1
and
nleB2
; however, their expression is independent of the LEE-encoded positive regulators Ler and GrlA but is repressed by GrlR and the global regulator H-NS. In contrast, GrlA and Ler are required for
nleA
expression, while H-NS silences it. Consistent with their role in the regulation of
nleA
, purified Ler and H-NS bound to the regulatory region of
nleA
upstream of its promoter. This work shows that at least two modes of regulation control the expression of effector genes in attaching and effacing (A/E) pathogens, suggesting that a subset of effector functions may be coordinately expressed in a particular niche or time during infection.
Functional Characterization of the Stringent Response Regulatory GenedksAof Vibrio cholerae and Its Role in Modulation of Virulence Phenotypes
Authors: Pal Ritesh Ranjan , Bag Satyabrata , Dasgupta Shreya , Das Bhabatosh , Bhadra Rupak K.
Published: August 2012

In bacteria, nutrient deprivation evokes the stringent response, which is mediated by the small intracellular signaling molecule ppGpp. In Gram negatives, the RelA enzyme synthesizes and SpoT hydrolyzes ppGpp, although the latter protein also has weak synthetase activity. DksA, a recently identified RNA polymerase binding transcription factor, acts as a coregulator along with ppGpp for controlling the stringent response. Recently, we have shown that three genes,relA,spoT, andrelV, govern cellular levels of ppGpp during various starvation stresses in the Gram-negative cholera pathogenVibrio cholerae. Here we report functional characterization of thedksAgene ofV. cholerae(dksAVc), coding for the protein DksAVc. Extensive genetic analyses of the ΔdksAVcmutants suggest that DksAVcis an important component involved in the stringent response inV. cholerae. Further analysis of mutants revealed that DksAVcpositively regulates various virulence-related processes, namely, motility, expression of the major secretory protease, called hemagglutinin protease (HAP), and production of cholera toxin (CT), underin vitroconditions. We found that DksAVcupregulates expression of the sigma factor FliA (σ28), a critical regulator of motility inV. cholerae. Altogether, it appears that apart from stringent-response regulation, DksAVcalso has important roles in fine regulation of virulence-related phenotypes ofV. cholerae.
Deletion of the Desulfovibrio vulgaris Carbon Monoxide Sensor Invokes Global Changes in Transcription
Authors: Rajeev Lara , Hillesland Kristina L. , Zane Grant M. , Zhou Aifen , Joachimiak Marcin P. , He Zhili , Zhou Jizhong , Arkin Adam P. , Wall Judy D. , Stahl David A.
Published: August 2012

The carbon monoxide-sensing transcriptional factor CooA has been studied only in hydrogenogenic organisms that can grow using CO as the sole source of energy. Homologs for the canonical CO oxidation system, including CooA, CO dehydrogenase (CODH), and a CO-dependent Coo hydrogenase, are present in the sulfate-reducing bacteriumDesulfovibrio vulgaris, although it grows only poorly on CO. We show thatD. vulgarisHildenborough has an active CO dehydrogenase capable of consuming exogenous CO and that the expression of the CO dehydrogenase, but not that of a gene annotated as encoding a Coo hydrogenase, is dependent on both CO and CooA. Carbon monoxide did not act as a general metabolic inhibitor, since growth of a strain deleted forcooAwas inhibited by CO on lactate-sulfate but not pyruvate-sulfate. While the deletion strain did not accumulate CO in excess, as would have been expected if CooA were important in the cycling of CO as a metabolic intermediate, global transcriptional analyses suggested that CooA and CODH are used during normal metabolism.
Direct and Indirect Regulation of the
ycnKJI
Operon Involved in Copper Uptake through Two Transcriptional Repressors, YcnK and CsoR, in Bacillus subtilis
Authors: Hirooka Kazutake , Edahiro Takayosh , Kimura Kosuke , Fujita Yasutaro
Published: August 2012

Northern blot and primer extension analyses revealed that the
ycnKJI
operon and the
ycnL
gene of
Bacillus subtilis
are transcribed from adjacent promoters that are divergently oriented. The
ycnK
and
ycnJ
genes encode a DeoR-type transcriptional regulator and a membrane protein involved in copper uptake, respectively. DNA binding experiments showed that the YcnK protein specifically binds to the
ycnK
-
ycnL
intergenic region, including a 16-bp direct repeat that is essential for the high binding affinity of YcnK, and that a copper-specific chelator significantly inhibits YcnK's DNA binding.
lacZ
reporter analysis showed that the
ycnK
promoter is induced by copper limitation or
ycnK
disruption. These results are consistent with YcnK functioning as a copper-responsive repressor that derepresses
ycnKJI
expression under copper limitation. On the other hand, the
ycnL
promoter was hardly induced by copper limitation, but
ycnK
disruption resulted in a slight induction of the
ycnL
promoter, suggesting that YcnK also represses
ycnL
weakly. Moreover, while the CsoR protein did not bind to the
ycnK
-
ycnL
intergenic region,
lacZ
reporter analysis demonstrated that
csoR
disruption induces the
ycnK
promoter only in the presence of intact
ycnK
and
copZA
genes. Since the
copZA
operon is involved in copper export and repressed by CsoR, it appears that the constitutive
copZA
expression brought by
csoR
disruption causes intracellular copper depletion, which releases the repression of the
ycnKJI
operon by YcnK.
Transcriptional Cross-Regulation between Gram-Negative and Gram-Positive Bacteria, Demonstrated Using ArgP-
argO
of Escherichia coli and LysG-
lysE
of Corynebacterium glutamicum
Authors: Marbaniang Carmelita N. , Gowrishankar J.
Published: August 2012

The protein-gene pairs ArgP-
argO
of
Escherichia coli
and LysG-
lysE
of
Corynebacterium glutamicum
are orthologous, with the first member of each pair being a LysR-type transcriptional regulator and the second its target gene encoding a basic amino acid exporter. Whereas LysE is an exporter of arginine (Arg) and lysine (Lys) whose expression is induced by Arg, Lys, or histidine (His), ArgO exports Arg alone, and its expression is activated by Arg but not Lys or His. We have now reconstituted in
E. coli
the activation of
lysE
by LysG in the presence of its coeffectors and have shown that neither ArgP nor LysG can regulate expression of the noncognate orthologous target. Of several ArgP-dominant (ArgP
d
) variants that confer elevated Arg-independent
argO
expression, some (ArgP
d
-P274S, -S94L, and, to a lesser extent, -P108S) activated
lysE
expression in
E. coli
. However, the individual activating effects of LysG and ArgP
d
on
lysE
were mutually extinguished when both proteins were coexpressed in Arg- or His-supplemented cultures. In comparison with native ArgP, the active ArgP
d
variants exhibited higher affinity of binding to the
lysE
regulatory region and less DNA bending at both
argO
and
lysE
. We conclude that the transcription factor LysG from a Gram-positive bacterium,
C. glutamicum
, is able to engage appropriately with the RNA polymerase from a Gram-negative bacterium,
E. coli
, for activation of its cognate target
lysE in vivo
and that single-amino-acid-substitution variants of ArgP can also activate the distantly orthologous target
lysE
, but by a subtly different mechanism that renders them noninterchangeable with LysG.
Polynucleotide Phosphorylase Plays an Important Role in the Generation of Spontaneous Mutations in Escherichia coli
Authors: Becket Elinne , Tse Lawrence , Yung Madeline , Cosico Alexander , Miller Jeffrey H.
Published: August 2012

Polynucleotide phosphorylase (PNP) plays a central role in RNA degradation, generating a pool of ribonucleoside diphosphates (rNDPs) that can be converted to deoxyribonucleoside diphosphates (dNDPs) by ribonucleotide reductase. We report here that spontaneous mutations resulting from replication errors, which are normally repaired by the mismatch repair (MMR) system, are sharply reduced in a PNP-deficient
Escherichia coli
strain. This is true for base substitution mutations that occur in the
rpoB
gene leading to Rif
r
and the
gyrB
gene leading to Nal
r
and for base substitution and frameshift mutations that occur in the
lacZ
gene. These results suggest that the increase in the rNDP pools generated by polynucleotide phosphorylase (PNP) degradation of RNA is responsible for the spontaneous mutations observed in an MMR-deficient background. The PNP-derived pool also appears responsible for the observed mutations in the
mutT
mutator background and those that occur after treatment with 5-bromodeoxyuridine, as these mutations are also drastically reduced in a PNP-deficient strain. However, mutation frequencies are not reduced in a
mutY
mutator background or after treatment with 2-aminopurine. These results highlight the central role in mutagenesis played by the rNDP pools (and the subsequent dNTP pools) derived from RNA degradation.
Mutational and Structural Analysis ofl-N-Carbamoylase Reveals New Insights into a Peptidase M20/M25/M40 Family Member
Authors: Martínez-Rodríguez Sergio , García-Pino Abel , Las Heras-Vázquez Francisco Javier , Clemente-Jiménez Josefa María , Rodríguez-Vico Felipe , García-Ruiz Juan M. , Loris Remy , Gavira Jose Antonio
Published: August 2012

N-Carbamoyl-l-amino acid amidohydrolases (l-carbamoylases) are important industrial enzymes used in kinetic resolution of racemic mixtures ofN-carbamoyl-amino acids due to their strict enantiospecificity. In this work, we report the firstl-carbamoylase structure belonging toGeobacillus stearothermophilusCECT43 (BsLcar), at a resolution of 2.7 Å. Structural analysis of BsLcar and several members of the peptidase M20/M25/M40 family confirmed the expected conserved residues at the active site in this family, and site-directed mutagenesis revealed their relevance to substrate binding. We also found an unexpectedly conserved arginine residue (Arg234in BsLcar), proven to be critical for dimerization of the enzyme. The mutation of this sole residue resulted in a total loss of activity and prevented the formation of the dimer in BsLcar. Comparative studies revealed that the dimerization domain of the peptidase M20/M25/M40 family is a “small-molecule binding domain,” allowing further evolutionary considerations for this enzyme family.
The Spanin Complex Is Essential for Lambda Lysis
Authors: Berry Joel , Rajaure Manoj , Pang Ting , Young Ry
Published: August 2012

Phage lysis is a ubiquitous biological process, the most frequent cytocidal event in the biosphere. Lysis of Gram-negative hosts has been shown to require holins and endolysins, which attack the cytoplasmic membrane and peptidoglycan, respectively. Recently, a third class of lysis proteins, the spanins, was identified. The first spanins to be characterized were λ Rz and Rz1, an integral cytoplasmic membrane protein and an outer membrane lipoprotein, respectively. Previous work has shown that Rz and Rz1 form complexes that span the entire periplasm. Phase-contrast video microscopy was used to record the morphological changes involved in the lysis of induced λ lysogens carrying prophages with either the λ canonical holin-endolysin system or the phage 21 pinholin-signal anchor release (SAR) endolysin system. In the former, rod morphology persisted until the instant of an explosive polar rupture, immediately emptying the cell of its contents. In contrast, in pinholin-SAR endolysin lysis, the cell began to shorten and thicken uniformly, with the resultant rounded cell finally bursting. In both cases, lysis failed to occur in inductions of isogenic prophages carrying null mutations in the spanin genes. In both systems, instead of an envelope rupture, the induced cells were converted from a rod shape to a spherical form. A functional GFPΦRz chimera was shown to exhibit a punctate distribution when coexpressed with Rz1, despite the absence of endolysin function. A model is proposed in which the spanins carry out the essential step of disrupting the outer membrane, in a manner regulated by the state of the peptidoglycan layer.
Functional Heterogeneity of the UpaH Autotransporter Protein from Uropathogenic Escherichia coli
Authors: Allsopp Luke P. , Beloin Christophe , Moriel Danilo Gomes , Totsika Makrina , Ghigo Jean-Marc , Schembri Mark A.
Published: August 2012

Uropathogenic
Escherichia coli
(UPEC) is responsible for the majority of urinary tract infections (UTI). To cause a UTI, UPEC must adhere to the epithelial cells of the urinary tract and overcome the shear flow forces of urine. This function is mediated primarily by fimbrial adhesins, which mediate specific attachment to host cell receptors. Another group of adhesins that contributes to UPEC-mediated UTI is autotransporter (AT) proteins. AT proteins possess a range of virulence properties, such as adherence, aggregation, invasion, and biofilm formation. One recently characterized AT protein of UPEC is UpaH, a large adhesin-involved-in-diffuse-adherence (AIDA-I)-type AT protein that contributes to biofilm formation and bladder colonization. In this study we characterized a series of naturally occurring variants of UpaH. We demonstrate that extensive sequence variation exists within the passenger-encoding domain of UpaH variants from different UPEC strains. This sequence variation is associated with functional heterogeneity with respect to the ability of UpaH to mediate biofilm formation. In contrast, all of the UpaH variants examined retained a conserved ability to mediate binding to extracellular matrix (ECM) proteins. Bioinformatic analysis of the UpaH passenger domain identified a conserved region (UpaH
CR
) and a hydrophobic region (UpaH
HR
). Deletion of these domains reduced biofilm formation but not the binding to ECM proteins. Despite variation in the
upaH
sequence, the transcription of
upaH
was repressed by a conserved mechanism involving the global regulator H-NS, and mutation of the
hns
gene relieved this repression. Overall, our findings shed new light on the regulation and functions of the UpaH AT protein.
Analysis of the Effects of a
gerP
Mutation on the Germination of Spores of Bacillus subtilis
Authors: Butzin Xuan Yi , Troiano Anthony J. , Coleman William H. , Griffiths Keren K. , Doona Christopher J. , Feeherry Florence E. , Wang Guiwen , Li Yong-qing , Setlow Peter
Published: August 2012

As previously reported,
gerP Bacillus subtilis
spores were defective in nutrient germination triggered via various germinant receptors (GRs), and the defect was eliminated by severe spore coat defects. The
gerP
spores' GR-dependent germination had a longer lag time between addition of germinants and initiation of rapid release of spores' dipicolinic acid (DPA), but times for release of >90% of DPA from individual spores were identical for wild-type and
gerP
spores. The
gerP
spores were also defective in GR-independent germination by DPA with its associated Ca
2+
divalent cation (CaDPA) but germinated better than wild-type spores with the GR-independent germinant dodecylamine. The
gerP
spores exhibited no increased sensitivity to hypochlorite, suggesting that these spores have no significant coat defect. Overexpression of GRs in
gerP
spores did lead to faster germination via the overexpressed GR, but this was still slower than germination of comparable
gerP
+
spores. Unlike wild-type spores, for which maximal nutrient germinant concentrations were between 500 μM and 2 mM for
l
-alanine and ≤10 mM for
l
-valine, rates of
gerP
spore germination increased up to between 200 mM and 1 M
l
-alanine and 100 mM
l
-valine, and at 1 M
l
-alanine, the rates of germination of wild-type and
gerP
spores with or without all alanine racemases were almost identical. A high pressure of 150 MPa that triggers spore germination by activating GRs also triggered germination of wild-type and
gerP
spores identically. All these results support the suggestion that GerP proteins facilitate access of nutrient germinants to their cognate GRs in spores' inner membrane.
Functional Analysis of the Genes Encoding Diaminopropionate Ammonia Lyase in Escherichia coli and Salmonella enterica Serovar Typhimurium
Authors: Kalyani J. N. , Ramachandra Nagaraju , Kachroo Aashiq H. , Mahadevan S. , Savithri H. S.
Published: August 2012

Diaminopropionate ammonia lyase (DAPAL) is a pyridoxal-5′phosphate (PLP)-dependent enzyme that catalyzes the conversion of diaminopropionate (DAP) to pyruvate and ammonia and plays an important role in cell metabolism. We have investigated the role of the
ygeX
gene of
Escherichia coli
K-12 and its ortholog,
STM1002
, in
Salmonella enterica
serovar Typhimurium LT2, presumed to encode DAPAL, in the growth kinetics of the bacteria. While
Salmonella
Typhimurium LT2 could grow on
dl-
DAP as a sole carbon source, the wild-type
E. coli
K-12 strain exhibited only marginal growth on
dl-DAP
, suggesting that DAPAL is functional in
S.
Typhimurium
.
The expression of
ygeX
in
E. coli
was low as detected by reverse transcriptase PCR (RT-PCR), consistent with the poor growth of
E. coli
on
dl-
DAP. Strains of
S.
Typhimurium and
E. coli
with
STM1002
and
ygeX
, respectively, deleted showed loss of growth on
dl-DAP
, confirming that
STM1002
(
ygeX
) is the locus encoding DAPAL. Interestingly, the presence of
dl-
DAP caused a growth inhibition of the wild-type
E. coli
strain as well as the knockout strains of
S.
Typhimurium and
E. coli
in minimal glucose/glycerol medium. Inhibition by
dl-
DAP was rescued by transforming the strains with plasmids containing the
STM1002
(
ygeX
) gene encoding DAPAL or supplementing the medium with Casamino Acids. Growth restoration studies using media lacking specific amino acid supplements suggested that growth inhibition by
dl-
DAP in the absence of DAPAL is associated with auxotrophy related to the inhibition of the enzymes involved in the biosynthetic pathways of pyruvate and aspartate and the amino acids derived from them.
Genomic Basis of a Polyagglutinating Isolate of Neisseria meningitidis
Authors: Rishishwar Lavanya , Katz Lee S. , Sharma Nitya V. , Rowe Lori , Frace Michael , Dolan Thomas Jennifer , Harcourt Brian H. , Mayer Leonard W. , Jordan I. King
Published: August 2012

Containment strategies for outbreaks of invasive
Neisseria meningitidis
disease are informed by serogroup assays that characterize the polysaccharide capsule. We sought to uncover the genomic basis of conflicting serogroup assay results for an isolate (M16917) from a patient with acute meningococcal disease. To this end, we characterized the complete genome sequence of the M16917 isolate and performed a variety of comparative sequence analyses against
N. meningitidis
reference genome sequences of known serogroups. Multilocus sequence typing and whole-genome sequence comparison revealed that M16917 is a member of the ST-11 sequence group, which is most often associated with serogroup C. However, sequence similarity comparisons and phylogenetic analysis showed that the serogroup diagnostic capsule polymerase gene (
synD
) of M16917 belongs to serogroup B. These results suggest that a capsule-switching event occurred based on homologous recombination at or around the capsule locus of M16917. Detailed analysis of this locus uncovered the locations of recombination breakpoints in the M16917 genome sequence, which led to the introduction of an ∼2-kb serogroup B sequence cassette into the serogroup C genomic background. Since there is no currently available vaccine for serogroup B strains of
N. meningitidis
, this kind capsule-switching event could have public health relevance as a vaccine escape mutant.
Meningoencephalitis with Subdural Empyema Caused by Toxigenic Clostridium perfringens Type A
Authors: Hugelshofer Michael , Achermann Yvonne , Kovari Helen , Dent Wolfgang , Hombach Michael , Bloemberg Guido
Published: August 2012

We report a clinical case of meningoencephalitis with subdural empyema in an immunocompromised farmer caused by toxigenic
Clostridium perfringens
type A, which was identified by 16S RNA gene analysis of cerebrospinal fluid and subdural empyema. In immunocompromised patients,
C. perfringens
should be considered a potential pathogen of sepsis.
Ampicillin Susceptibility Can Predict
In Vitro
Susceptibility of Penicillin-Resistant, Ampicillin-Susceptible Enterococcus faecalis Isolates to Amoxicillin but Not to Imipenem and Piperacillin
Authors: Conceição Natália , de Oliveira Cristina da Cunha Hueb Barata , da Silva Lucas Emanuel Pinheiro , de Souza Laís Rezende Cardoso , de Oliveira Adriana Gonçalves
Published: August 2012

Our findings demonstrated that the results obtained for ampicillin may accurately predict the
in vitro
susceptibility to amoxicillin but not to imipenem and piperacillin among isolates of
Enterococcus faecalis
resistant to penicillin but susceptible to ampicillin, which have emerged recently, in contrast to penicillin- and ampicillin-susceptible isolates.
Peptide Interactions Stabilize and Restructure Human Papillomavirus Type 16 E6 To Interact with p53
Authors: Ansari Tina , Brimer Nicole , Vande Pol Scott B.
Published: August 2012

Human papillomavirus type 16 (HPV-16) E6 (16E6) binds the E3 ubiquitin ligase E6AP and p53, thereby targeting degradation of p53 (M. Scheffner, B. A. Werness, J. M. Huibregtse, A. J. Levine, and P. M. Howley, Cell 63:1129–1136, 1990). Here we show that minimal 16E6-binding LXXLL peptides reshape 16E6 to confer p53 interaction and stabilize 16E6
in vivo
but that degradation of p53 by 16E6 requires E6AP expression. These experiments establish a general mechanism for how papillomavirus E6 binding to LXXLL peptides reshapes E6 to then act as an adapter molecule.
A Three-Helix Junction Is the Interface between Two Functional Domains of Prohead RNA in ϕ29 DNA Packaging
Authors: Zhao Wei , Saha Mitul , Ke Ailong , Morais Marc C. , Jardine Paul J. , Grimes Shelley
Published: August 2012

The double-stranded-DNA bacteriophages employ powerful molecular motors to translocate genomic DNA into preformed capsids during the packaging step in phage assembly.
Bacillus subtilis
bacteriophage ϕ29 has an oligomeric prohead RNA (pRNA) that is an essential component of its packaging motor. The crystal structure of the pRNA-prohead binding domain suggested that a three-helix junction constitutes both a flexible region and part of a rigid RNA superhelix. Here we define the functional role of the three-helix junction in motor assembly and DNA packaging. Deletion mutagenesis showed that a U-rich region comprising two sides of the junction plays a role in the stable assembly of pRNA to the prohead. The retention of at least two bulged residues in this region was essential for pRNA binding and thereby subsequent DNA packaging. Additional deletions resulted in the loss of the ability of pRNA to multimerize in solution, consistent with the hypothesis that this region provides the flexibility required for pRNA oligomerization and prohead binding. The third side of the junction is part of a large RNA superhelix that spans the motor. The insertion of bases into this feature resulted in a loss of DNA packaging and an impairment of initiation complex assembly. Additionally, cryo-electron microscopy (cryoEM) analysis of third-side insertion mutants showed an increased flexibility of the helix that binds the ATPase, suggesting that the rigidity of the RNA superhelix is necessary for efficient motor assembly and function. These results highlight the critical role of the three-way junction in bridging the prohead binding and ATPase assembly functions of pRNA.
The Cytoplasmic Tail Domain of Influenza B Virus Hemagglutinin Is Important for Its Incorporation into Virions but Is Not Essential for Virus Replication in Cell Culture in the Presence of Compensatory Mutations
Authors: Imai Masaki , Watanabe Shinji , Kawaoka Yoshihiro
Published: August 2012

Influenza B virus hemagglutinin (BHA) contains a predicted cytoplasmic tail of 10 amino acids that are highly conserved among influenza B viruses. To understand the role of this cytoplasmic tail in infectious virus production, we used reverse genetics to generate a recombinant influenza B virus lacking the BHA cytoplasmic tail domain. The resulting virus, designated BHATail
−
, had a titer approximately 5 log units lower than that of wild-type virus but grew normally when BHA was supplemented in
trans
by BHA-expressing cells. Although the levels of BHA cell surface expression were indistinguishable between truncated and wild-type BHA, the BHATail
−
virus produced particles containing dramatically less BHA. Moreover, removal of the cytoplasmic tail abrogated the association of BHA with Triton X-100-insoluble lipid rafts. Interestingly, long-term culture of a virus lacking the BHA cytoplasmic tail in Madin-Darby canine kidney (MDCK) cells yielded a mutant with infectivities somewhat similar to that of wild-type virus. Sequencing revealed that the mutant virus retained the original cytoplasmic tail deletion but acquired additional mutations in its BHA, neuraminidase (NA), and M1 proteins. Viral growth kinetic analysis showed that replication of BHA cytoplasmic tailless viruses could be improved by compensatory mutations in the NA and M1 proteins. These findings indicate that the cytoplasmic tail domain of BHA is important for efficient incorporation of BHA into virions and tight lipid raft association. They also demonstrate that the domain is not absolutely required for virus viability in cell culture in the presence of compensatory mutations.
Factors influencing infant sleep position: decisions do not differ by SES in African-American families
Authors: Robida David , Moon Rachel Y
Published: August 2012

Objective

To investigate factors influencing African-American parents’ knowledge, attitudes and practice regarding infant sleep position and determine if these differ by socioeconomic status (SES).

Methods

A cross-sectional sample of 412 parents with infants ≤6 months of age participated in a validated survey of knowledge, attitudes and practice.

Results

There was no significant difference in attitudes or practice, and knowledge was similar regarding infant sleep position between African-American parents of higher and lower SES. The healthcare provider recommendation of exclusive supine sleep position use was associated with increased knowledge, overall decreased use of the side position (5.0% vs 16.8%, p<0.01) and increased occasional use of the supine position in the lower SES group (81.6% vs 68.6%, p=0.03). It was not associated with increased positive parental attitudes about the supine sleep position in either group. Neither a senior caregiver living in the home nor observation of hospital personnel placing infants in a non-supine position was associated with differences in sleep position practices in either group.

Conclusions

Sleep position practices in African-American families do not differ by SES. Improved attitudes toward positioning and increased use of supine positioning may result if healthcare providers address common concerns and misconceptions about sleep position.
Low injury rate strongly correlates with team success in Qatari professional football
Authors: Eirale Cristiano , Tol J L , Farooq Abdulaziz , Smiley Faten , Chalabi Hakim
Published: August 2012

Background

Although the incidence of football injuries should relate to team success there is little empirical evidence.

Objective

We investigated the relationship between injury incidence and team success in Qatar first-division football clubs.

Methods

Using a prospective cohort study design, we captured exposure and injuries in Qatar male elite football for a season. Club performance was measured by total league points, ranking, goal scored, goals conceded and number of matches won, drawn or lost.

Results

Lower injury incidence was strongly correlated with team ranking position (r=0.929, p=0.003), more games won (r=0.883, p=0.008), more goals scored (r=0.893, p=0.007), greater goal difference (r=0.821, p=0.003) and total points (r=0.929, p=0.003).

Conclusions

Lower incidence rate was strongly correlated with team success. Prevention of injuries may contribute to team success.
Upper limb injury in rugby union football: results of a cohort study
Authors: Usman Juliana , McIntosh Andrew Stuart
Published: August 2012

Background

There have been few in-depth studies of upper limb injury epidemiology in rugby union football, despite reports that they accounted for between 14% and 28% of all rugby injuries.

Objective

To report on upper limb injury incidence, injury severity and to identify the risk factors associated with upper limb injuries, for example, level of play, season (years) and playing position.

Design

Prospective cohort study across five rugby seasons from 2004 to 2008.

Setting

Formal rugby competitions—suburban, provincial and international.

Participants

1475 adult male rugby players in Colts, Grade and Elite competitions.

Main outcome measurements

An upper limb injury resulting in a missed game and its characteristics.

Results

A total of 61 598 athletic exposures (AE) and 606 upper limb injuries were recorded. About 66% of the injuries were to the shoulder. The overall upper limb injury incidence rate (IIR) was 9.84 injuries/1000 AE (95% CI 9.06 to 10.62). Statistically significant associations were found between upper limb injuries and level of play; and between shoulder injuries and playing position (p<0.05). No association was found between upper limb and shoulder injuries and study year. The overall upper limb IIR decreased as the level of play increased; 10.74 upper limb injuries/1000 AE (95% CI 9.93 to 11.56) in Colts to 6.07 upper limb injuries/1000 AE (95% CI 5.46 to 6.69) in Elite.

Conclusions

The upper limb IIR decreased as the level of play increased indicating that age, level of skill and playing experience may be risk factors for upper limb injury.
Do pregnant women follow exercise guidelines? Prevalence data among 3482 women, and prediction of low-back pain, pelvic girdle pain and depression
Authors: Gjestland Kristin , Bø Kari , Owe Katrine Mari , Eberhard-Gran Malin
Published: August 2012

Purpose

We describe exercise level in mid-pregnancy, associated sociodemographic variables, and investigate the association between exercise in mid-pregnancy and subsequent low-back pain, pelvic girdle pain and depression at 32 weeks of pregnancy.

Material and methods

The study included 3482 pregnant women participating in the Akershus Birth Cohort study (response rate 80.5%). Data were collected by a questionnaire in pregnancy weeks 17–21, pregnancy week 32 and electronic birth journal. The results were analysed by logistic regression and are presented as crude (cOR) and adjusted OR (aOR) with 95% CI.

Results

Only 14.6% of the respondents followed the current exercise prescription for exercise during pregnancy (≥3 times a week, >20 min at moderate intensity). One-third of the study sample exercised less than once a week at pregnancy weeks 17–21. Women exercising either 1–2 times or ≥3 times a week at mid-pregnancy were more often primiparous, higher-educated and had less often prepregnacy body mass index >30 kg/m2compared with women exercising less than once a week. Women who exercised ≥3 times a week were less likely to report pelvic girdle pain (aOR: 0.76, 95% CI 0.61 to 0.96), while women exercising 1–2 times a week were less likely to report low-back pain (aOR: 0.80, 95% CI 0.66 to 0.97) and depression (aOR: 0.66, 95% CI 0.48 to 0.91).

Conclusions

Few Norwegian women follow current exercise prescriptions for exercise in mid-pregnancy. The results may indicate an association between exercising mid-pregnancy and lower prevalence of low-back pain, pelvic girdle pain and depression in late pregnancy.
ICAM-1K469E polymorphism is a genetic determinant for the clinical risk factors of T2D subjects with retinopathy in Indians: a population-based case–control study
Authors: Vinita Kumari , Sripriya Sarangapani , Prathiba Krishnamurthy , Vaitheeswaran Kulothungan , Sathyabaarathi Ravichandran , Rajesh Mahendran , Amali John , Umashankar Vetrivel , Kumaramanickavel Govindasamy , Pal Swakshyar Saumya , Raman Rajiv , Sharma Tarun
Published: August 2012

Objective

Elevated levels of intercellular adhesion molecule-1 (ICAM-1) are demonstrated in diabetes complications. The current study aims to understand association of K469E (rs5498) inICAM-1gene, in type 2 diabetic (T2D) subjects with retinopathy.

Design

Case–control study.

Setting

Sankara Nethralaya Diabetic Retinopathy Epidemiology and Molecular Genetic Study, an epidemiology study (on prevalence of diabetic retinopathy in T2D subjects (T2DR) from south India) and outpatient department of Sankara Nethralaya, a tertiary care hospital, in Chennai, India.

Participants

A total of 356 T2D subjects of >15 years of diabetes duration, with (n=199) and without (n=157) retinopathy.

Methods

The rs5498 polymorphism was genotyped by direct sequencing. Multivariate analysis for various clinical covariates was done using SPSS V.14. Comparative assessment of structure stability, folding rate of the variants were assessed using bioinformatics tools like STRIDE, MuPro, ModellerV97, fold rate server, etc.

Results

The AA genotype of rs5498 was seen at a higher frequency in the retinopathy group (p=0.012). The risk for diabetic retinopathy (DR) increased in the presence of AA genotype (OR=1.89–4.82) after the sequential addition of various clinical covariates. Multivariate logistic regression analysis showed 8.26 times high risk for developing DR in the AG genotype (p=0.003). Structural superimposition of ICAM-1 wild type (K469) and variant (E469) showed 0.943 Å of backbone root mean square deviation as calculated by PYMOL software. A difference in the fold rate time was also observed between the wild type (5.4/s) and variant (3.3/s).

Conclusions

This study shows that allele A of rs5498 inICAM-1is a putative risk predisposing allele for T2D retinopathy and its clinical covariates in Indian population. The folding rate of the protein decreases for the A allele implicating a potential effect on the structure and function of ICAM-1.
A randomised cross-over cohort study of exposure to emissions from a road tunnel ventilation stack
Authors: Cowie Christine T , Ezz Wafaa , Xuan Wei , Lilley William , Rose Nectarios , Rae Michael , Marks Guy B
Published: August 2012

Background and objective

Road tunnels are increasingly important components of urban infrastructure. However, knowledge of their health impact on surrounding communities is limited. Our objective was to estimate the short-term respiratory health effects of exposure to emissions from a road tunnel ventilation stack.

Methods

We conducted a randomised cross-over cohort study in 36 volunteers who underwent three exposure scenarios in 2006 before the road tunnel opened, and in 2007 (n=27) and 2008 (n=20) after the tunnel opened. Exposure downwind of the stack was compared to upwind of the stack and to a distant heavily trafficked location adjacent to a main road. Spirometry, exhaled nitric oxide (eNO) and symptom scores were measured repeatedly during each 2 h exposure session.

Results

Downwind locations were associated with increased reports of ‘dry nose’ (score difference 0.36; 95% CI 0.09 to 0.63) compared with the control location (2006 vs 2007/2008), but not with impaired lung function, increased airway inflammation or other symptoms. The heavily trafficked location was associated with significantly increased eNO (ratio=1.09; 95% CI 1.04 to 1.14), eye (score difference 0.05; 95% CI 0.01 to 0.10) and chest (score difference 0.21; 95% CI 0.09 to 0.33) symptoms compared to the stack locations.

Conclusions

There was no consistent evidence of adverse respiratory effects from short-term exposures downwind of the tunnel ventilation stack, except for dry nose symptoms. However, the findings of increased airway inflammation and symptoms in subjects after only 2 h exposure at the heavily trafficked location, are suggestive of detrimental effects of short-term exposures to traffic-related air pollution.
What components of chronic care organisation relate to better primary care for coronary heart disease patients? An observational study
Authors: van Lieshout Jan , Frigola Capell Eva , Ludt Sabine , Grol Richard , Wensing Michel
Published: August 2012

Objectives

Cardiovascular risk management (CVRM) received by patients shows large variation across countries. In this study we explored the aspects of primary care organisation associated with key components of CVRM in coronary heart disease (CHD) patients.

Design

Observational study.

Setting

273 primary care practices in Austria, Belgium, England, Finland, France, Germany, The Netherlands, Slovenia, Switzerland and Spain.

Participants

A random sample of 4563 CHD patients identified by coded diagnoses in eight countries, based on prescription lists and while visiting the practice in one country each.

Main outcome measure

We performed an audit in primary care practices in 10 European countries. We used six indicators to measure key components of CVRM: risk factor recording, antiplatelet therapy, influenza vaccination, blood pressure levels (systolic <140 and diastolic <90 mm Hg), and low-density lipoprotein cholesterol <2.5 mmol/l. Data from structured questionnaires were used to construct an overall measure and six domain measures of practice organisation based on 39 items. Using multilevel regression analyses we explored the effects of practice organisation on CVRM, controlling for patient characteristics.

Results

Better overall organisation of a primary care practice was associated with higher scores on three indicators: risk factor registration (B=0.0307, p<0.0001), antiplatelet therapy (OR 1.05, p=0.0245) and influenza vaccination (OR 1.12, p<0.0001). Overall practice organisation was not found to be related with recorded blood pressure or cholesterol levels. Only the organisational domains ‘self-management support’ and ‘use of clinical information systems’ were linked to three CVRM indicators.

Conclusions

A better organisation of a primary care practice was associated with better scores on process indicators of CVRM in CHD patients, but not on intermediate patient outcome measures. Direct support for patients and clinicians seemed most influential.
Two-dimensional strain profiles in patients with physiological and pathological hypertrophy and preserved left ventricular systolic function: a comparative analyses
Authors: Afonso Luis , Kondur Ashok , Simegn Mengistu , Niraj Ashutosh , Hari Pawan , Kaur Ramanjit , Ramappa Preeti , Pradhan Jyotiranjan , Bhandare Deepti , Williams Kim A , Zalawadiya Sandip , Pinheiro Aurelio , Abraham Theodore P
Published: August 2012

Objective

This study was designed to examine the utility of two-dimensional strain (2DS) or speckle tracking imaging to typify functional adaptations of the left ventricle in variant forms of left ventricular hypertrophy (LVH).

Design

Cross-sectional study.

Setting

Urban tertiary care academic medical centres.

Participants

A total of 129 subjects, 56 with hypertrophic cardiomyopathy (HCM), 34 with hypertensive left ventricular hypertrophy (H-LVH), 27 professional athletes with LVH (AT-LVH) and 12 healthy controls in sinus rhythm with preserved left ventricular systolic function.

Methods

Conventional echocardiographic and tissue Doppler examinations were performed in all study subjects. Bi-dimensional acquisitions were analysed to map longitudinal systolic strain (automated function imaging, AFI, GE Healthcare, Waukesha, Wisconsin, USA) from apical views.

Results

Subjects with HCM had significantly lower regional and average global peak longitudinal systolic strain (GLS-avg) compared with controls and other forms of LVH. Strain dispersion index, a measure of regional contractile heterogeneity, was higher in HCM compared with the rest of the groups. On receiver operator characteristics analysis, GLS-avg had excellent discriminatory ability to distinguish HCM from H-LVH area under curve (AUC) (0.893, p<0.001) or AT-LVH AUC (0.920, p<0.001). Tissue Doppler and LV morphological parameters were better suited to differentiate the athlete heart from HCM.

Conclusions

2DS (AFI) allows rapid characterisation of regional and global systolic function and may have the potential to differentiate HCM from variant forms of LVH.
Impact of television coverage on the number and type of symptoms reported during a health scare: a retrospective pre–post observational study
Authors: Faasse Kate , Gamble Greg , Cundy Tim , Petrie Keith J
Published: August 2012

Objectives

This study investigated the impact of television news coverage on total adverse event reporting rates 1 month before and after the bulletins during a medication health scare. We further investigated whether individual side effects mentioned in each bulletin were reflected in the adverse event reports following the coverage.

Design

A retrospective pre–post observational study.

Setting

New Zealand Centre for Adverse Reactions Monitoring.

Participants

Adverse events reported from May to December 2008 relating to Eltroxin formulation change.

Primary and secondary outcome measures

Primary outcome measure was the total rate of adverse event reporting per day. Secondary outcome measure was the rate of reporting of seven individual symptoms mentioned in the television coverage.

Results

After story 1, a significant increase in total reporting rates was evident (MdnPre=0, MdnPost=13.5, U=2, p<0.001, r=−0.86) with larger effect sizes for increases in television-mentioned symptoms. Story 2 also showed a significant increase in total adverse event reporting (MdnPre=6, MdnPost=18.5, U=86.5, p=0.002, r=−0.49) driven by significant increases only in television-reported symptoms. Story 3 did not result in a significant increase in total reporting (MdnPre=12; MdnPost=15.5, U=171, p=0.432, r=−0.12), and showed a significant increase in reporting rates for only one of the two television-reported symptoms.

Conclusions

The findings suggest that television news coverage can impact on the overall rate of adverse event reporting during a health scare, in part via increased reporting of media-mentioned side effects. The effects of television media coverage on adverse event reporting appear strongest for earlier reports.
Prehospital tracheal intubation in severe facial trauma: a short window of opportunity
Authors: P P Bredmose , S Rock , D J Lockey
Published: August 2012

A case of severe facial injury is described. The pre-hospital management including pre-hospital anaesthesia and intubation and the importance of advanced anaesthetic skills in rare trauma cases is discussed. In addition the rare situation where large bone fragments are retrieved from the scene and potentially used in reconstruction is mentioned.
A new typology of policies to tackle health inequalities and scenarios of impact based on Rose's population approach
Authors: Benach Joan , Malmusi Davide , Yasui Yutaka , Martínez José Miguel
Published: August 2012

The last decade has witnessed a surge in interest for policies to tackle health inequalities. Adequate theoretical development of policy models is needed to understand how to design and evaluate equity-oriented health policies. In this paper we review Graham's typology of policies (focused on the worst-off, on the gap, or on the gradient) and propose an adaptation (targeted, universal with additional targeting, redistributive, and proportionate universalism). For each type, potential scenarios of impact on population health and health inequalities are depicted following the idea of Geoffrey Rose's population curves and strategies, policy examples are given and a simulation with survey data is shown. The proposed typology of scenarios of health inequality reduction can serve as an effective tool to interpret the differential impact of interventions and to reflect on how to adequately design or re-orient a policy and which measures to use to evaluate it.
Severe venous congestive encephalopathy secondary to a dialysis arteriovenous graft
Authors: Samaniego Edgar A , Abrams Kevin J , Dabus Guilherme , Starr Rosanne , Linfante Italo
Published: August 2012

The clinical presentation and imaging of venous congestive encephalopathy (VCE) can mimic several neurological conditions, making the diagnosis challenging. We report a patient with end stage renal disease on dialysis who presented with a right occipital infarction. The patient developed progressive encephalopathy and increased intracranial pressure. Extensive imaging, electroencephalography and serum analysis did not explain the cause of his infarction and progressive neurological deterioration. Finally, cerebral angiography and venography demonstrated severe generalized VCE due to arterial shunting from a right upper extremity arteriovenous graft (AVG) and an occluded right innominate venous trunk. The right arm shunt resulted in severe cerebral venous hypertension due to ipsilateral occlusion of the innominate venous trunk. After the AVG was repaired, the cerebral venous hypertension resolved and the patient returned to baseline.