bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of a Pyridoxine-Derived Small-Molecule Inhibitor Targeting Dengue Virus RNA-Dependent RNA Polymerase
Authors: Xu Hong-Tao , Colby-Germinario Susan P. , Hassounah Said , Quashie Peter K. , Han Yingshan , Oliveira Maureen , Stranix Brent R. , Wainberg Mark A. 
Published: November 2015

ABSTRACT

The viral RNA-dependent RNA polymerase (RdRp) activity of the dengue virus (DENV) NS5 protein is an attractive target for drug design. Here, we report the identification of a novel class of inhibitor (i.e., an active-site metal ion chelator) that acts against DENV RdRp activity. DENV RdRp utilizes a two-metal-ion mechanism of catalysis; therefore, we constructed a small library of compounds, through mechanism-based drug design, aimed at chelating divalent metal ions in the catalytic site of DENV RdRp. We now describe a pyridoxine-derived small-molecule inhibitor that targets DENV RdRp and show that 5-benzenesulfonylmethyl-3-hydroxy-4-hydroxymethyl-pyridine-2-carboxylic acid hydroxyamide (termed DMB220) inhibited the RdRp activity of DENV serotypes 1 to 4 at low micromolar 50% inhibitory concentrations (IC
50s
of 5 to 6.7 μM) in an enzymatic assay. The antiviral activity of DMB220 against DENV infection was also verified in a cell-based assay and showed a 50% effective concentration (EC
50
) of <3 μM. Enzyme assays proved that DMB220 was competitive with nucleotide incorporation. DMB220 did not inhibit the enzymatic activity of recombinant HIV-1 reverse transcriptase and showed only weak inhibition of HIV-1 integrase strand transfer activity, indicating high specificity for DENV RdRp. S600T substitution in the DENV RdRp, which was previously shown to confer resistance to nucleoside analogue inhibitors (NI), conferred 3-fold hypersusceptibility to DMB220, and enzymatic analyses showed that this hypersusceptibility may arise from the decreased binding/incorporation efficiency of the natural NTP substrate without significantly impacting inhibitor binding. Thus, metal ion chelation at the active site of DENV RdRp represents a viable anti-DENV strategy, and DMB220 is the first of a new class of DENV inhibitor.
Inhibition of Pseudomonas aeruginosa ExsA DNA-Binding Activity by
N
-Hydroxybenzimidazoles
Authors: Marsden Anne E. , King Jessica M. , Spies M. Ashley , Kim Oak K. , Yahr Timothy L. 
Published: November 2015

ABSTRACT

The
Pseudomonas aeruginosa
type III secretion system (T3SS) is a primary virulence determinant and a potential target for antivirulence drugs. One candidate target is ExsA, a member of the AraC family of DNA-binding proteins required for expression of the T3SS. A previous study identified small molecules based on an
N
-hydroxybenzimidazole scaffold that inhibit the DNA-binding activity of several AraC proteins, including ExsA. In this study, we further characterized a panel of
N
-hydroxybenzimidazoles. The half-maximal inhibitory concentrations (IC
50
s) for the tested
N
-hydroxybenzimidazoles ranged from 8 to 45 μM in DNA-binding assays. Each of the
N
-hydroxybenzimidazoles protected mammalian cells from T3SS-dependent cytotoxicity, and protection correlated with reduced T3SS gene expression in a coculture infection model. Binding studies with the purified ExsA DNA-binding domain (i.e., lacking the amino-terminal self-association domain) confirmed that the activity of
N
-hydroxybenzimidazoles results from interactions with the DNA-binding domain. The interaction is specific, as an unrelated DNA-binding protein (Vfr) was unaffected by
N
-hydroxybenzimidazoles. ExsA homologs that control T3SS gene expression in
Yersinia pestis
,
Aeromonas hydrophila
, and
Vibrio parahaemolyticus
were also sensitive to
N
-hydroxybenzimidazoles. Although ExsA and
Y. pestis
LcrF share 79% sequence identity in the DNA-binding domain, differential sensitivities to several of the
N
-hydroxybenzimidazoles were observed. Site-directed mutagenesis based on
in silico
docking of inhibitors to the DNA-binding domain, and on amino acid differences between ExsA and LcrF, resulted in the identification of several substitutions that altered the sensitivity of ExsA to
N
-hydroxybenzimidazoles. Development of second-generation compounds targeted to the same binding pocket could lead to drugs with improved pharmacological properties.
Whole-Genome Sequencing in Microbial Forensic Analysis of Gamma-Irradiated Microbial Materials
Authors: Broomall Stacey M. , Ait Ichou Mohamed , Krepps Michael D. , Johnsky Lauren A. , Karavis Mark A. , Hubbard Kyle S. , Insalaco Joseph M. , Betters Janet L. , Redmond Brady W. , Rivers Bryan A. , Liem Alvin T. , Hill Jessica M. , Fochler Edward T. , Roth Pierce A. , Rosenzweig C. Nicole , Skowronski Evan W. , Gibbons Henry S. 
Published: November 2015

ABSTRACT
Effective microbial forensic analysis of materials used in a potential biological attack requires robust methods of morphological and genetic characterization of the attack materials in order to enable the attribution of the materials to potential sources and to exclude other potential sources. The genetic homogeneity and potential intersample variability of many of the category A to C bioterrorism agents offer a particular challenge to the generation of attributive signatures, potentially requiring whole-genome or proteomic approaches to be utilized. Currently, irradiation of mail is standard practice at several government facilities judged to be at particularly high risk. Thus, initial forensic signatures would need to be recovered from inactivated (nonviable) material. In the study described in this report, we determined the effects of high-dose gamma irradiation on forensic markers of bacterial biothreat agent surrogate organisms with a particular emphasis on the suitability of genomic DNA (gDNA) recovered from such sources as a template for whole-genome analysis. While irradiation of spores and vegetative cells affected the retention of Gram and spore stains and sheared gDNA into small fragments, we found that irradiated material could be utilized to generate accurate whole-genome sequence data on the Illumina and Roche 454 sequencing platforms.
Characterization of the Vibrio fischeri Fatty Acid Chemoreceptors, VfcB and VfcB2
Authors: Nikolakakis K. , Monfils K. , Moriano-Gutierrez S. , Brennan C. A. , Ruby E. G. 
Published: November 2015

ABSTRACT

Bacteria use a wide variety of methyl-accepting chemotaxis proteins (MCPs) to mediate their attraction to or repulsion from different chemical signals in their environment. The bioluminescent marine bacterium
Vibrio fischeri
is the monospecific symbiont of the Hawaiian bobtail squid,
Euprymna scolopes
, and encodes a large repertoire of MCPs that are hypothesized to be used during different parts of its complex, multistage lifestyle. Here, we report the initial characterization of two such MCPs from
V. fischeri
that are responsible for mediating migration toward short- and medium-chain aliphatic (or fatty) acids. These receptors appear to be distributed among only members of the family
Vibrionaceae
and are likely descended from a receptor that has been lost by the majority of the members of this family. While chemotaxis greatly enhances the efficiency of host colonization by
V. fischeri
, fatty acids do not appear to be used as a chemical cue during this stage of the symbiosis. This study presents an example of straight-chain fatty acid chemoattraction and contributes to the growing body of characterized MCP-ligand interactions.
Bromochloromethane, a Methane Analogue, Affects the Microbiota and Metabolic Profiles of the Rat Gastrointestinal Tract
Authors: Yang Yu-Xiang , Mu Chun-Long , Luo Zhen , Zhu Wei-Yun 
Published: November 2015

ABSTRACT
Bromochloromethane (BCM), an inhibitor of methanogenesis, has been used in animal production. However, little is known about its impact on the intestinal microbiota and metabolic patterns. The present study aimed to investigate the effect of BCM on the colonic bacterial community and metabolism by establishing a Wistar rat model. Twenty male Wistar rats were randomly divided into two groups (control and treated with BCM) and raised for 6 weeks. Bacterial fermentation products in the cecum were determined, and colonic methanogens and sulfate-reducing bacteria (SRB) were quantified. The colonic microbiota was analyzed by pyrosequencing of the 16S rRNA genes, and metabolites were profiled by gas chromatography and mass spectrometry. The results showed that BCM did not affect body weight and feed intake, but it did significantly change the intestinal metabolic profiles. Cecal protein fermentation was enhanced by BCM, as methylamine, putrescine, phenylethylamine, tyramine, and skatole were significantly increased. Colonic fatty acid and carbohydrate concentrations were significantly decreased, indicating the perturbation of lipid and carbohydrate metabolism by BCM. BCM treatment decreased the abundance of methanogen populations, while SRB were increased in the colon. BCM did not affect the total colonic bacterial counts but significantly altered the bacterial community composition by decreasing the abundance of actinobacteria, acidobacteria, and proteobacteria. The results demonstrated that BCM treatment significantly altered the microbiotic and metabolite profiles in the intestines, which may provide further information on the use of BCM in animal production.
Targeted Amplicon Sequencing for Single-Nucleotide-Polymorphism Genotyping of Attaching and Effacing Escherichia coli O26:H11 Cattle Strains via a High-Throughput Library Preparation Technique
Authors: Ison Sarah A. , Delannoy Sabine , Bugarel Marie , Nagaraja Tiruvoor G. , Renter David G. , den Bakker Henk C. , Nightingale Kendra K. , Fach Patrick , Loneragan Guy H. 
Published: November 2015

ABSTRACT

Enterohemorrhagic
Escherichia coli
(EHEC) O26:H11, a serotype within Shiga toxin-producing
E. coli
(STEC) that causes severe human disease, has been considered to have evolved from attaching and effacing
E. coli
(AEEC) O26:H11 through the acquisition of a Shiga toxin-encoding gene. Targeted amplicon sequencing using next-generation sequencing technology of 48 phylogenetically informative single-nucleotide polymorphisms (SNPs) and three SNPs differentiating Shiga toxin-positive (
stx
-positive) strains from Shiga toxin-negative (
stx
-negative) strains were used to infer the phylogenetic relationships of 178
E. coli
O26:H11 strains (6
stx
-positive strains and 172
stx
-negative AEEC strains) from cattle feces to 7 publically available genomes of human clinical strains. The AEEC cattle strains displayed synonymous SNP genotypes with
stx
2
-positive sequence type 29 (ST29) human O26:H11 strains, while
stx
1
ST21 human and cattle strains clustered separately, demonstrating the close phylogenetic relatedness of these Shiga toxin-negative AEEC cattle strains and human clinical strains. With the exception of seven
stx
-negative strains, five of which contained
espK
, three
stx
-related SNPs differentiated the STEC strains from non-STEC strains, supporting the hypothesis that these AEEC cattle strains could serve as a potential reservoir for new or existing pathogenic human strains. Our results support the idea that targeted amplicon sequencing for SNP genotyping expedites strain identification and genetic characterization of
E. coli
O26:H11, which is important for food safety and public health.
Influence of IgG Subclass on Human Antimannan Antibody-Mediated Resistance to Hematogenously Disseminated Candidiasis in Mice
Authors: Nishiya Casey T. , Boxx Gayle M. , Robison Kerry , Itatani Carol , Kozel Thomas R. , Zhang Mason X. 
Published: November 2015

ABSTRACTCandida albicansis a yeast-like pathogen and can cause life-threatening systemic candidiasis. Its cell surface is enriched with mannan that is resistant to complement activation. Previously, we developed the recombinant human IgG1 antimannan antibody M1g1. M1g1 was found to promote complement activation and phagocytosis and protect mice from systemic candidiasis. Here, we evaluate the influence of IgG subclass on antimannan antibody-mediated protection. Three IgG subclass variants of M1g1 were constructed: M1g2, M1g3, and M1g4. The IgG subclass identity for each variant was confirmed with DNA sequence and subclass-specific antibodies. These variants contain identical M1 Fabs and exhibited similar binding affinities forC. albicansyeast and purified mannan. Yeast cells and hyphae recovered from the kidney of antibody-treated mice with systemic candidiasis showed uniform binding of each variant, indicating constitutive expression of the M1 epitope and antibody opsonization in the kidney. All variants promoted deposition of both murine and human C3 onto the yeast cell surface, with M1g4 showing delayed activation, as determined by flow cytometry and immunofluorescence microscopy. M1g4-mediated complement activation was found to be associated with its M1 Fab that activates the alternative pathway in an Fc-independent manner. Treatment with each subclass variant extended the survival of mice with systemic candidiasis (P< 0.001). However, treatment with M1g1, M1g3, or M1g4, but not with M1g2, also reduced the kidney fungal burden (P< 0.001). Thus, the role of human antimannan antibody in host resistance to systemic candidiasis is influenced by its IgG subclass.
Host Genetic Variations and Sex Differences Potentiate Predisposition, Severity, and Outcomes of Group A Streptococcus-Mediated Necrotizing Soft Tissue Infections
Authors: Chella Krishnan Karthickeyan , Mukundan Santhosh , Alagarsamy Jeyashree , Laturnus Donna , Kotb Malak 
Published: November 2015

ABSTRACT

Host genetic variations play an important role in several pathogenic diseases, and we previously provided strong evidence that these genetic variations contribute significantly to differences in susceptibility and clinical outcomes of invasive group A
Streptococcus
(GAS) patients, including sepsis and necrotizing soft tissue infections (NSTIs). The goal of the present study was to investigate how genetic variations and sex differences among four commonly used mouse strains contribute to variation in severity, manifestations, and outcomes of NSTIs. DBA/2J mice were more susceptible to NSTIs than C57BL/6J, BALB/c, and CD-1 mice, as exhibited by significantly greater bacteremia, excessive dissemination to the spleen, and significantly higher mortality. Differences in the sex of the mice also contributed to differences in disease severity and outcomes: DBA/2J female mice were relatively resistant compared to their male counterparts. However, DBA/2J mice exhibited minimal weight loss and developed smaller lesions than did the aforementioned strains. Moreover, at 48 h after infection, compared with C57BL/6J mice, DBA/2J mice had increased bacteremia, excessive dissemination to the spleen, and excessive concentrations of inflammatory cytokines and chemokines. These results indicate that variations in the host genetic context as well as sex play a dominant role in determining the severity of and susceptibility to GAS NSTIs.
2-Aminoacrylate Stress Induces a Context-Dependent Glycine Requirement in
ridA
Strains of Salmonella enterica
Authors: Ernst Dustin C. , Downs Diana M. 
Published: November 2015

ABSTRACT

The reactive enamine 2-aminoacrylate (2AA) is a metabolic stressor capable of damaging cellular components. Members of the broadly conserved Rid (RidA/YER057c/UK114) protein family mitigate 2AA stress
in vivo
by facilitating enamine and/or imine hydrolysis. Previous work showed that 2AA accumulation in
ridA
strains of
Salmonella enterica
led to the inactivation of multiple target enzymes, including serine hydroxymethyltransferase (GlyA). However, the specific cause of a
ridA
strain's inability to grow during periods of 2AA stress had yet to be determined. Work presented here shows that glycine supplementation suppressed all 2AA-dependent
ridA
strain growth defects described to date. Depending on the metabolic context, glycine appeared to suppress
ridA
strain growth defects by eliciting a GcvB small RNA-dependent regulatory response or by serving as a precursor to one-carbon units produced by the glycine cleavage complex (GCV). In either case, the data suggest that GlyA is the most physiologically sensitive target of 2AA inactivation in
S. enterica
. The universally conserved nature of GlyA among free-living organisms highlights the importance of RidA in mitigating 2AA stress.

IMPORTANCE
The RidA stress response prevents 2-aminoacrylate (2AA) damage from occurring in prokaryotes and eukaryotes alike. 2AA inactivation of serine hydroxymethyltransferase (GlyA) from
Salmonella enterica
restricts glycine and one-carbon production, ultimately reducing fitness of the organism. The cooccurrence of genes encoding 2AA production enzymes and serine hydroxy-methyltransferase (SHMT) in many genomes may in part underlie the evolutionary selection for Rid proteins to maintain appropriate glycine and one-carbon metabolism throughout life.
Biochemical Characterization of the Flagellar Rod Components of Rhodobacter sphaeroides: Properties and Interactions
Authors: Osorio-Valeriano Manuel , de la Mora Javier , Camarena Laura , Dreyfus Georges 
Published: November 2015

ABSTRACT

The flagellar basal body is a rotary motor that spans the cytoplasmic and outer membranes. The rod is a drive shaft that transmits torque generated by the motor through the hook to the filament that propels the bacterial cell. The assembly and structure of the rod are poorly understood. In a first attempt to characterize this structure in the alphaproteobacterium
Rhodobacter sphaeroides
, we overexpressed and purified FliE and the four related rod proteins (FlgB, FlgC, FlgF, and FlgG), and we analyzed their ability to form homo-oligomers. We found that highly purified preparations of these proteins formed high-molecular-mass oligomers that tended to dissociate in the presence of NaCl. As predicted by
in silico
modeling, the four rod proteins share architectural features. Using affinity blotting, we detected the heteromeric interactions between these proteins. In addition, we observed that deletion of the N- and C-terminal regions of FlgF and FlgG severely affected heteromeric but not homomeric interactions. On the basis of our findings, we propose a model of rod assembly in this bacterium.

IMPORTANCE
Despite the considerable amount of research on the structure and assembly of other flagellar axial structures that has been conducted, the rod has been barely studied. An analysis of the biochemical characteristics of the flagellar rod components of the Fla1 system of
R. sphaeroides
is presented in this work. We also analyze the interactions of these proteins with each other and with their neighbors, and we propose a model for the order in which they are assembled.
Rethinking the Hierarchy of Sugar Utilization in Bacteria
Authors: Beisel Chase L. , Afroz Taliman 
Published: November 2015

ABSTRACTBacteria are known to consume some sugars over others, although recent work reported by Koirala and colleagues in this issue of theJournal of Bacteriology(S. Koirala, X. Wang, and C. V. Rao, J Bacteriol 198:386–393, 2016,http://dx.doi.org/10.1128/JB.00709-15) revealed that individual cells do not necessarily follow this hierarchy. By studying the preferential consumption ofl-arabinose overd-xylose inEscherichia coli, those authors found that subpopulations consume one, the other, or both sugars through cross-repression between utilization pathways. Their findings challenge classic assertions about established hierarchies and can guide efforts to engineer the simultaneous utilization of multiple sugars.
The efficacy of extraembryonic stem cells in improving blood flow within animal models of lower limb ischaemia
Authors: Mohamed Omer Safraz , Krishna Smriti Murali , Li Jiaze , Moxon Joseph Vaughan , Nsengiyumva Vianne , Golledge Jonathan 
Published: November 2015

BackgroundStem cell (SC) administration is a potential therapeutic strategy to improve blood supply in patients with peripheral artery disease (PAD). The aim of this systematic review and meta-analysis was to investigate the efficacy of extraembryonic tissue-derived SC (ETSC) in improving blood flow within animal models of hindlimb ischaemia (HLI).MethodsPubMed, ScienceDirect and Web of Science were searched to identify studies which investigated ETSCs within animal HLI models. A meta-analysis was performed focusing on the effect of ETSCs on limb blood flow assessed by laser Doppler imaging using a random effects model. Methodological quality was assessed using a newly devised quality assessment tool.ResultsFive studies investigating umbilical cord-derived SCs (three studies), placental SCs (one study), amnion and chorionic SCs (one study) were included. A meta-analysis suggested that administration of ETSCs improved the restoration of blood flow within the HLI models used. The methodological quality of the included studies was assessed as poor. Problems identified included lack of randomised design and blinding of outcome assessors; that the animal models did not incorporate recognised risk factors for human PAD or atherosclerosis; the models used did not have established chronic ischaemia as is the cases in most patients presenting with PAD; and the studies lacked a clear rationale for the dosage and frequency of SCs administered.ConclusionsThe identified studies suggest that ETSCs improve recovery of limb blood supply within current animal HLI models. Improved study quality is, however, needed to provide support for the likelihood of translating these findings to patients with PAD.
‘First, do no harm’: are disability assessments associated with adverse trends in mental health? A longitudinal ecological study
Authors: Barr B , Taylor-Robinson D , Stuckler D , Loopstra R , Reeves A , Whitehead M 
Published: November 2015

BackgroundIn England between 2010 and 2013, just over one million recipients of the main out-of-work disability benefit had their eligibility reassessed using a new functional checklist—the Work Capability Assessment. Doctors and disability rights organisations have raised concerns that this has had an adverse effect on the mental health of claimants, but there are no population level studies exploring the health effects of this or similar policies.MethodWe used multivariable regression to investigate whether variation in the trend in reassessments in each of 149 local authorities in England was associated with differences in local trends in suicides, self-reported mental health problems and antidepressant prescribing rates, while adjusting for baseline conditions and trends in other factors known to influence mental ill-health.ResultsEach additional 10 000 people reassessed in each area was associated with an additional 6 suicides (95% CI 2 to 9), 2700 cases of reported mental health problems (95% CI 548 to 4840), and the prescribing of an additional 7020 antidepressant items (95% CI 3930 to 10100). The reassessment process was associated with the greatest increases in these adverse mental health outcomes in the most deprived areas of the country, widening health inequalities.ConclusionsThe programme of reassessing people on disability benefits using the Work Capability Assessment was independently associated with an increase in suicides, self-reported mental health problems and antidepressant prescribing. This policy may have had serious adverse consequences for mental health in England, which could outweigh any benefits that arise from moving people off disability benefits.