bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation of Benznidazole by Trypanosomal Type I Nitroreductases Results in Glyoxal Formation
Authors: Hall Belinda S. , Wilkinson Shane R.
Published: October 2011

Benznidazole, a 2-nitroimidazole, is the front-line treatment used against American trypanosomiasis, a parasitic infection caused by
Trypanosoma cruzi
. Despite nearly 40 years of use, the trypanocidal activity of this prodrug is not fully understood. It has been proposed that benznidazole activation leads to the formation of reductive metabolites that can cause a series of deleterious effects, including DNA damage and thiol depletion. Here, we show that the key step in benznidazole activation involves an NADH-dependent trypanosomal type I nitroreductase. This catalyzes an oxygen-insensitive reaction with the interaction of enzyme, reductant, and prodrug occurring through a ping-pong mechanism. Liquid chromatography/mass spectrometry (LC/MS) analysis of the resultant metabolites identified 4,5-dihydro-4,5-dihydroxyimidazole as the major product of a reductive pathway proceeding through hydroxylamine and hydroxy intermediates. The breakdown of this product released the reactive dialdehyde glyoxal, which, in the presence of guanosine, generated guanosine-glyoxal adducts. These experiments indicate that the reduction of benznidazole by type I nitroreductase activity leads to the formation of highly reactive metabolites and that the expression of this enzyme is key to the trypanocidal properties displayed by the prodrug.
An Enlarged, Adaptable Active Site in CYP164 Family P450 Enzymes, the Sole P450 in Mycobacterium leprae
Authors: Agnew Christopher R. J. , Warrilow Andrew G. S. , Burton Nicholas M. , Lamb David C. , Kelly Steven L. , Brady R. Leo
Published: October 2011

CYP164 family P450 enzymes are found in only a subset of mycobacteria and include CYP164A1, which is the sole P450 found in
Mycobacterium leprae
, the causative agent of leprosy. This has previously led to interest in this enzyme as a potential drug target. Here we describe the first crystal structure of a CYP164 enzyme, CYP164A2 from
Mycobacterium smegmatis
. CYP164A2 has a distinctive, enlarged hydrophobic active site that extends above the porphyrin ring toward the access channels. Unusually, we find that CYP164A2 can simultaneously bind two econazole molecules in different regions of the enlarged active site and is accompanied by the rearrangement and ordering of the BC loop. The primary location is through a classic interaction of the azole group with the porphyrin iron. The second econazole molecule is bound to a unique site and is linked to a tetracoordinated metal ion complexed to one of the heme carboxylates and to the side chains of His 105 and His 364. All of these features are preserved in the closely homologous
M. leprae
CYP164A1. The computational docking of azole compounds to a homology model of CYP164A1 suggests that these compounds will form effective inhibitors and is supported by the correlation of parallel docking with experimental binding studies of CYP164A2. The binding of econazole to CYP164A2 occurs primarily through the high-spin “open” conformation of the enzyme (

K
d

[dissociation constant] of 0.1 μM), with binding to the low-spin “closed” form being significantly hindered (

K
d

of 338 μM). These studies support previous suggestions that azole derivatives may provide an effective strategy to improve the treatment of leprosy.
Molecular Dynamics Approaches Estimate the Binding Energy of HIV-1 Integrase Inhibitors and Correlate with
In Vitro
Activity
Authors: Johnson Barry C. , Métifiot Mathieu , Pommier Yves , Hughes Stephen H.
Published: October 2011

The design of novel integrase (IN) inhibitors has been aided by recent crystal structures revealing the binding mode of these compounds with a full-length prototype foamy virus (PFV) IN and synthetic viral DNA ends. Earlier docking studies relied on incomplete structures and did not include the contribution of the viral DNA to inhibitor binding. Using the structure of PFV IN as the starting point, we generated a model of the corresponding HIV-1 complex and developed a molecular dynamics (MD)-based approach that correlates with the
in vitro
activities of novel compounds. Four well-characterized compounds (raltegravir, elvitegravir, MK-0536, and dolutegravir) were used as a training set, and the data for their
in vitro
activity against the Y143R, N155H, and G140S/Q148H mutants were used in addition to the wild-type (WT) IN data. Three additional compounds were docked into the IN-DNA complex model and subjected to MD simulations. All three gave interaction potentials within 1 standard deviation of values estimated from the training set, and the most active compound was identified. Additional MD analysis of the raltegravir- and dolutegravir-bound complexes gave internal and interaction energy values that closely match the experimental binding energy of a compound related to raltegravir that has similar activity. These approaches can be used to gain a deeper understanding of the interactions of the inhibitors with the HIV-1 intasome and to identify promising scaffolds for novel integrase inhibitors, in particular, compounds that retain activity against a range of drug-resistant mutants, making it possible to streamline synthesis and testing.
In Vitro
and
In Vivo
Trypanocidal Synergistic Activity of
N
-Butyl-1-(4-Dimethylamino)Phenyl-1,2,3,4-Tetrahydro-β-Carboline-3-Carboxamide Associated with Benznidazole
Authors: Valdez Rodrigo Hinojosa , Tonin Lilian Tatiani Düsman , Ueda-Nakamura Tânia , Silva Sueli Oliveira , Dias Filho Benedito Prado , Kaneshima Edilson Nobuyoshi , Yamada-Ogatta Sueli Fumie , Yamauchi Lucy Megumi , Sarragiotto Maria Helena , Nakamura Celso Vataru
Published: October 2011

American trypanosomiasis, or Chagas' disease, is caused by
Trypanosoma cruzi
and affects around 15 million people throughout the American continent. The available treatment is based on two nitroheterocyclic drugs, nifurtimox and benznidazole, both only partially effective and toxic. In this context, new drugs must be found. In our previous work, the tetrahydro-β-carboline compound
N
-butyl-1-(4-dimethylamino)phenyl-1,2,3,4-tetrahydro-β-carboline-3-carboxamide, named C4, showed a potent
in vitro
trypanocidal effect. The goal of this study was to evaluate the
in vitro
and
in vivo
trypanocidal effects of the compound C4 associated with other drugs (benznidazole, ketoconazole, and amphotericin B). For this, we used the checkerboard technique to analyze the effect of combinations of C4 reference drugs. C4 was assayed in a murine model alone as well as in association with benznidazole. We also evaluated the parasitemia, mortality, weight, and presence of amastigote nests in cardiac tissue. A synergic effect of C4 plus benznidazole against epimastigote and trypomastigote forms was observed
in vitro
, and in the murine model, we observed a substantial reduction in parasitemia levels and lowered mortality rates. These findings encourage supplementary investigations of carboline compounds as potential new trypanocidal drugs.
Bacteriocin Production: a Probiotic Trait?
Authors: Dobson Alleson , Cotter Paul D. , Ross R. Paul , Hill Colin
Published: October 2011

Bacteriocins are an abundant and diverse group of ribosomally synthesized antimicrobial peptides produced by bacteria and archaea. Traditionally, bacteriocin production has been considered an important trait in the selection of probiotic strains, but until recently, few studies have definitively demonstrated the impact of bacteriocin production on the ability of a strain to compete within complex microbial communities and/or positively influence the health of the host. Although research in this area is still in its infancy, there is intriguing evidence to suggest that bacteriocins may function in a number of ways within the gastrointestinal tract. Bacteriocins may facilitate the introduction of a producer into an established niche, directly inhibit the invasion of competing strains or pathogens, or modulate the composition of the microbiota and influence the host immune system. Here we review the role of bacteriocin production in complex microbial communities and their potential to enhance human health.
Genome Sequences and Characterization of the Related Gordonia Phages GTE5 and GRU1 and Their Use as Potential Biocontrol Agents
Authors: Petrovski Steve , Tillett Daniel , Seviour Robert J.
Published: October 2011

Activated sludge plants suffer frequently from the operational problem of stable foam formation on aerobic reactor surfaces, which can be difficult to prevent. Many foams are stabilized by mycolic acid-containing
Actinobacteria
, the mycolata. The
in situ
biocontrol of foaming using phages is an attractive strategy. We describe two polyvalent phages, GTE5 and GRU1, targeting
Gordonia terrae
and
Gordonia rubrupertincta
, respectively, isolated from activated sludge. Phage GRU1 also propagates on
Nocardia nova
. Both phages belong to the family
Siphoviridae
and have similar-size icosahedral heads that encapsulate double-stranded DNA genomes (∼65 kb). Their genome sequences are similar to each other but markedly different from those of other sequenced phages. Both are arranged in a modular fashion. These phages can reduce or eliminate foam formation by their host cells under laboratory conditions.
High-Sensitivity Stable-Isotope Probing by a Quantitative Terminal Restriction Fragment Length Polymorphism Protocol
Authors: Andeer Peter , Strand Stuart E. , Stahl David A.
Published: October 2011

Stable-isotope probing (SIP) has proved a valuable cultivation-independent tool for linking specific microbial populations to selected functions in various natural and engineered systems. However, application of SIP to microbial populations with relatively minor buoyant density increases, such as populations that utilize compounds as a nitrogen source, results in reduced resolution of labeled populations. We therefore developed a tandem quantitative PCR (qPCR)–TRFLP (terminal restriction fragment length polymorphism) protocol that improves resolution of detection by quantifying specific taxonomic groups in gradient fractions. This method combines well-controlled amplification with TRFLP analysis to quantify relative taxon abundance in amplicon pools of FAM-labeled PCR products, using the intercalating dye EvaGreen to monitor amplification. Method accuracy was evaluated using mixtures of cloned 16S rRNA genes, DNA extracted from low- and high-G+C bacterial isolates (
Escherichia coli
,
Rhodococcus
,
Variovorax
, and
Microbacterium
), and DNA from soil microcosms amended with known amounts of genomic DNA from bacterial isolates. Improved resolution of minor shifts in buoyant density relative to TRFLP analysis alone was confirmed using well-controlled SIP analyses.
Endosymbiont Transmission Mode in Bacterial Leaf Nodulation as Revealed by a Population Genetic Study of Psychotria leptophylla
Authors: Lemaire Benny , Janssens Steven , Smets Erik , Dessein Steven
Published: October 2011

Leaf-nodulated plants are colonized by vertically inherited bacterial endosymbionts, which maintain symbioses throughout host generations. The permanent character of the interaction implies phylogenetic congruence between the host and the endosymbiont. However, the present population genetic study of
Psychotria leptophylla
provides evidence for a mixed symbiont transmission involving both vertical inheritance and horizontal transfers from the environment.
Iron Transformations Induced by an Acid-Tolerant Desulfosporosinus Species
Authors: Bertel Doug , Peck John , Quick Thomas J. , Senko John M.
Published: October 2011

The mineralogical transformations of Fe phases induced by an acid-tolerant, Fe(III)- and sulfate-reducing bacterium,Desulfosporosinussp. strain GBSRB4.2 were evaluated under geochemical conditions associated with acid mine drainage-impacted systems (i.e., low pH and high Fe concentrations). X-ray powder diffractometry coupled with magnetic analysis by first-order reversal curve diagrams were used to evaluate mineral phases produced by GBSRB4.2 in media containing different ratios of Fe(II) and Fe(III). In medium containing Fe predominately in the +II oxidation state, ferrimagnetic, single-domain greigite (Fe3S4) was formed, but the addition of Fe(III) inhibited greigite formation. In media that contained abundant Fe(III) [as schwertmannite; Fe8O8(OH)6SO4·nH2O], the activities of strain GBSRB4.2 enhanced the transformation of schwertmannite to goethite (α-FeOOH), due to the increased pH and Fe(II) concentrations that resulted from the activities of GBSRB4.2.
Comparison of Assays for Sensitive and Reproducible Detection of Cell Culture-Infectious Cryptosporidium parvum and Cryptosporidium hominis in Drinking Water
Authors: Johnson Anne M. , Di Giovanni George D. , Rochelle Paul A.
Published: October 2011

This study compared the three most commonly used assays for detecting
Cryptosporidium
sp. infections in cell culture: immunofluorescent antibody and microscopy assay (IFA), PCR targeting
Cryptosporidium
sp.-specific DNA, and reverse transcriptase PCR (RT-PCR) targeting
Cryptosporidium
sp.-specific mRNA. Monolayers of HCT-8 cells, grown in 8-well chamber slides or 96-well plates, were inoculated with a variety of viable and inactivated oocysts to assess assay performance. All assays detected infection with low doses of flow cytometry-enumerated
Cryptosporidium parvum
oocysts, including infection with one oocyst and three oocysts. All methods also detected infection with
Cryptosporidium hominis
. The RT-PCR assay, IFA, and PCR assay detected infection in 23%, 25%, and 51% of monolayers inoculated with three
C. parvum
oocysts and 10%, 9%, and 16% of monolayers inoculated with one oocyst, respectively. The PCR assay was the most sensitive, but it had the highest frequency of false positives with mock-infected cells and inactivated oocysts. IFA was the only infection detection assay that did not produce false positives with mock-infected monolayers. IFA was also the only assay that detected infections in all experiments with spiked oocysts recovered from Envirochek capsules following filtration of 1,000 liters of treated water. Consequently, cell culture with IFA detection is the most appropriate method for routine and sensitive detection of infectious
Cryptosporidium parvum
and
Cryptosporidium hominis
in drinking water.
Prevalence of Ca
2+
-ATPase-Mediated Carbonate Dissolution among Cyanobacterial Euendoliths
Authors: Ramírez-Reinat E. L. , Garcia-Pichel F.
Published: October 2011

Recent physiological work has shown that the filamentous euendolithic cyanobacterium
Mastigocoleus testarum
(strain BC008) is able to bore into solid carbonates using Ca
2+
-ATPases to take up Ca
2+
from the medium at the excavation front, promoting dissolution of CaCO
3
there. It is not known, however, if this is a widespread mechanism or, rather, a unique capability of this model strain. To test this, we undertook a survey of multispecies euendolithic microbial assemblages infesting natural carbonate substrates in marine coastal waters of the Caribbean, Mediterranean, South Pacific, and Sea of Cortez. Microscopic examination revealed the presence of complex assemblages of euendoliths, encompassing 3 out of the 5 major cyanobacterial orders. 16S rRNA gene clone libraries detected even greater diversity, particularly among the thin-filamentous forms, and allowed us to categorize the endoliths in our samples into 8 distinct phylogenetic clades. Using real-time Ca
2+
imaging under a confocal laser scanning microscope, we could show that all communities displayed light-dependent formation of Ca
2+
-supersaturated zones in and around boreholes, a staple of actively boring phototrophs. In 3 out of 4 samples, boring activity was sensitive to at least one of two inhibitors of Ca
2+
-ATPase transporters (thapsigargin or
tert
-butylhydroquinone), indicating that the Ca
2+
-ATPase mechanism is widespread among cyanobacterial euendoliths but perhaps not universal. Function-community structure correlations point to one particular clade of baeocyte-forming euendoliths as the potential exception.
Engineering of Bacterial Methyl Ketone Synthesis for Biofuels
Authors: Goh Ee-Been , Baidoo Edward E. K. , Keasling Jay D. , Beller Harry R.
Published: October 2011

We have engineered
Escherichia coli
to overproduce saturated and monounsaturated aliphatic methyl ketones in the C
11
to C
15
(diesel) range; this group of methyl ketones includes 2-undecanone and 2-tridecanone, which are of importance to the flavor and fragrance industry and also have favorable cetane numbers (as we report here). We describe specific improvements that resulted in a 700-fold enhancement in methyl ketone titer relative to that of a fatty acid-overproducing
E. coli
strain, including the following: (i) overproduction of β-ketoacyl coenzyme A (CoA) thioesters achieved by modification of the β-oxidation pathway (specifically, overexpression of a heterologous acyl-CoA oxidase and native FadB and chromosomal deletion of
fadA
) and (ii) overexpression of a native thioesterase (FadM). FadM was previously associated with oleic acid degradation, not methyl ketone synthesis, but outperformed a recently identified methyl ketone synthase (
Solanum habrochaites
MKS2 [
Sh
MKS2], a thioesterase from wild tomato) in β-ketoacyl-CoA-overproducing strains tested. Whole-genome transcriptional (microarray) studies led to the discovery that FadM is a valuable catalyst for enhancing methyl ketone production. The use of a two-phase system with decane enhanced methyl ketone production by 4- to 7-fold in addition to increases from genetic modifications.
Adhesive Bond Stiffness of Staphylococcus aureus with and without Proteins That Bind to an Adsorbed Fibronectin Film
Authors: Olsson Adam L. J. , Sharma Prashant K. , van der Mei Henny C. , Busscher Henk J.
Published: October 2011

Staphylococcus aureus
is known to cause biomaterial-associated infections of implants and devices once it has breached the skin and mucosal barriers. Adhesion is the initial step in the development of a biomaterial-associated infection, and strategies to prevent staphylococcal adhesion and thus biomaterial-associated infections require understanding of the adhesive bond. The aim of this study was to compare the adhesive bond stiffnesses of two
S. aureus
strains with and without fibronectin-binding proteins (FnBPs) adhering to a fibronectin-coated quartz crystal microbalance (QCM) sensor surface on the basis of a coupled- resonance model. Both fibronectin adsorption and staphylococcal adhesion were accompanied by negative frequency shifts, regardless of the absence or presence of FnBPs on the staphylococcal cell surfaces. This is the opposite of the positive frequency shifts often observed for other bacterial strains adhering to bare sensor surfaces. Most likely, adhering staphylococci sink into and deform the adsorbed protein layer, creating stiff binding with the sensor surface due to an increased bacterium-substratum contact area.
S. aureus
8325-4 possesses FnBPs and yields less negative frequency shifts (Δ
f
) that are further away from the zero-crossing frequency than
S. aureus
DU5883. This suggests that FnBPs on
S. aureus
8325-4 create a stiffer bond to the fibronectin coating than has been observed for
S. aureus
DU5883. Due to a limited window of observation, as defined by the available resonance frequencies in QCM, we could not determine exact stiffness values.
Protective Effects of Passively Transferred Merozoite-Specific Antibodies against Theileria equi in Horses with Severe Combined Immunodeficiency
Authors: Mealey Robert H. , Kappmeyer Lowell S. , Ueti Massaro W. , Wagner Bettina , Knowles Donald P.
Published: October 2011

Theileria equiimmune plasma was infused into young horses (foals) with severe combined immunodeficiency. Although all foals became infected following intravenous challenge with homologousT. equimerozoite stabilate, delayed time to peak parasitemia occurred. Protective effects were associated with a predominance of passively transferred merozoite-specific IgG3.
Serum Antibody Responses to 10 Mycobacterium tuberculosis Proteins, Purified Protein Derivative, and Old Tuberculin in Natural and Experimental Tuberculosis in Rhesus Monkeys
Authors: Min Fangui , Zhang Yu , Huang Ren , Li Wende , Wu Yu'e , Pan Jinchun , Zhao Weibo , Liu Xiangmei
Published: October 2011

Old tuberculin (OT) and purified protein derivative (PPD) are widely used for tuberculin skin testing (TST) in diagnosis of tuberculosis (TB) but often yield poor specificity and anergy in reaction. Therefore, it is necessary to develop new serological methods as a possible auxiliary diagnostic method for TB. In this study, we characterized the dynamic antibody responses of 10 purified recombinant antigens, PPD, and OT in rhesus monkeys experimentally infected with
Mycobacterium tuberculosis
and analyzed the time to antibody detection, antibody levels, and their association with the infectious doses. The antibodies were detected as early as 4 weeks after infection in response to 5 antigens (CFP10, CFP10-ESAT-6, U1, MPT64, and Ag85b). Antibodies against most of the other antigens were detected between 4 and 12 weeks after infection. The levels of antibodies were dose dependant. We further evaluated the serodiagnostic potential of these antigens by using indirect enzyme-linked immunosorbent assay in 71 TST-positive and 90 TST-negative serum samples from monkeys. For all 12 antigens, the median optical density values of TST-positive monkeys were statistically significantly higher than those of TST-negative monkeys (
P
< 0.001). Among those antigens, Ag85b and CFP10 showed higher diagnostic potential than others. A combination of results from Ag85b, the 38-kDa antigen (Ag38kDa), and Ag14kDa reaches a sensitivity of 95.77%, indicating that these antigens may be ideal cocktails in TB diagnosis.
Maintenance of Immune Response throughout Childhood following Serogroup C Meningococcal Conjugate Vaccination in Early Childhood
Authors: Khatami A. , Peters A. , Robinson H. , Williams N. , Thompson A. , Findlow H. , Pollard A. J. , Snape M. D.
Published: October 2011

The objectives of this study were to evaluate the kinetics of antibody decline through childhood in a longitudinal study of a single cohort following serogroup C meningococcal (MenC) vaccine immunization in early childhood and to calculate the proportion of 11 to 13 year olds with protective levels of bactericidal antibody 10 years after immunization. United Kingdom children aged 11 to 13 years in 2010 who had previously taken part in a longitudinal study at the Oxford Vaccine Group had blood samples drawn between 2001 and 2010. Sera from each time point were analyzed for the MenC serum bactericidal antibody titer using a baby rabbit complement (rSBA) assay. The median age at MenC immunization was 21 months (range, 1 year 3 months to 3 years 9 months). The MenC rSBA geometric mean titer (GMT) at age 3.5 to 5 years was 8.0 (95% confidence interval, 6.5 to 9.9;
n
= 287). By age 11.5 to 13.5 years, the rSBA GMT had declined to 3.3 (2.5 to 4.4;
n
= 98). The percentage of children with rSBA titers of ≥1:8 (the threshold for protection) also declined from 38% (35% to 41%) to 15% (12% to 19%). We concluded that MenC rSBA titers wane rapidly following vaccination in early childhood and continue to decline into the second decade of life. Since nasopharyngeal colonization in adolescents probably provides the major reservoir for MenC in the population, declining immunity in this cohort is of concern. Sustaining high levels of antibody through booster vaccination in this cohort is likely necessary to avoid a resurgence of disease in the decade ahead.
Structural Correlates of Cytoplasmic and Chloroplast Lipid Body Synthesis in Chlamydomonas reinhardtii and Stimulation of Lipid Body Production with Acetate Boost
Authors: Goodson Carrie , Roth Robyn , Wang Zi Teng , Goodenough Ursula
Published: October 2011

Light microscopy and deep-etch electron microscopy were used to visualize triacylglyceride (TAG)-filled lipid bodies (LBs) of the green eukaryotic soil alga
Chlamydomonas reinhardtii
, a model organism for biodiesel production. Cells growing in nitrogen-replete media contain small cytoplasmic lipid bodies (α-cyto-LBs) and small chloroplast plastoglobules. When starved for N, β-cyto-LB formation is massively stimulated. β-Cyto-LBs are intimately associated with both the endoplasmic reticulum membrane and the outer membrane of the chloroplast envelope, suggesting a model for the active participation of both organelles in β-cyto-LB biosynthesis and packaging. When
sta6
mutant cells, blocked in starch biosynthesis, are N starved, they produce β-cyto-LBs and also chloroplast LBs (cpst-LBs) that are at least 10 times larger than plastoglobules and eventually engorge the chloroplast stroma. Production of β-cyto-LBs and cpst-LBs under the conditions we used is dependent on exogenous 20 mM acetate. We propose that the greater TAG yields reported for N-starved
sta6
cells can be attributed to the strain's ability to produce cpst-LBs, a capacity that is lost when the mutant is complemented by a
STA6
transgene. Provision of a 20 mM acetate “boost” during N starvation generates
sta6
cells that become so engorged with LBs—at the expense of cytoplasm and most organelles—that they float on water even when centrifuged. This property could be a desirable feature for algal harvesting during biodiesel production.
The Intact
dupA
Cluster Is a More Reliable Helicobacter pylori Virulence Marker than
dupA
Alone
Authors: Jung Sung Woo , Sugimoto Mitsushige , Shiota Seiji , Graham David Y. , Yamaoka Yoshio
Published: October 2011

The duodenal ulcer promoting (
dupA
) gene, located in the plasticity region of
Helicobacter pylori
, is associated with duodenal ulcer development.
dupA
was predicted to form a type IV secretory system (T4SS) with
vir
genes around
dupA
(
dupA
cluster). We investigated the prevalence of
dupA
and
dupA
clusters and clarified associations between the
dupA
cluster status and clinical outcomes in the U.S. population. In all, 245
H. pylori
strains were examined using PCR to evaluate the status of
dupA
and the adjacent
vir
genes predicted to form T4SS, in addition to the status of
cag
pathogenicity island (PAI). The associations between
dupA
cluster status and interleukin-8 (IL-8) and IL-12 production were also examined. The presence of
dupA
and all adjacent
vir
genes were defined as a complete
dupA
cluster. Many variations related to the status of
dupA
and
dupA
cluster genes were identified. Concurrent
H. pylori
infection and the presence of a complete
dupA
cluster increases duodenal ulcer risk compared to
H. pylori
infection with incomplete
dupA
cluster or without the
dupA
gene independent on the
cag
PAI status (adjusted odds ratio, 2.13; 95% confidence interval, 1.13 to 4.03). Gastric mucosal IL-8 levels were also significantly higher in the complete
dupA
cluster group than in other groups (
P
= 0.01). In conclusion, although the causal relationship between the
dupA
cluster and duodenal ulcer development is not proved, the presence of a complete
dupA
cluster but not
dupA
alone, is associated with duodenal ulcer development.
LipC (Rv0220) Is an Immunogenic Cell Surface Esterase of Mycobacterium tuberculosis
Authors: Shen Guomiao , Singh Krishna , Chandra Dinesh , Serveau-Avesque Carole , Maurin Damien , Canaan Stéphane , Singla Rupak , Behera Digambar , Laal Suman
Published: October 2011

We have reported previously the identification of novel proteins of
Mycobacterium tuberculosis
by the immunoscreening of an expression library of
M. tuberculosis
genomic DNA with sera obtained from
M. tuberculosis
-infected rabbits at 5 weeks postinfection. In this study, we report the further characterization of one of these antigens, LipC (Rv0220). LipC is annotated as a member of the Lip family based on the presence of the consensus motif “GXSXG” characteristic of esterases. Although predicted to be a cytoplasmic enzyme, we provide evidence that LipC is a cell surface protein that is present in both the cell wall and the capsule of
M. tuberculosis
. Consistent with this localization, LipC elicits strong humoral immune responses in both HIV-negative (HIV
−
) and HIV-positive (HIV
+
) tuberculosis (TB) patients. The absence of anti-LipC antibodies in sera from purified protein derivative-positive (PPD
+
) healthy subjects confirms its expression only during active
M. tuberculosis
infection. Epitope mapping of LipC identified 6 immunodominant epitopes, 5 of which map to the exposed surface of the modeled LipC protein. The recombinant LipC (rLipC) protein also elicits proinflammatory cytokine and chemokine responses from macrophages and pulmonary epithelial cells. rLipC can hydrolyze short-chain esters with the carbon chain containing 2 to 10 carbon atoms. Together, these studies demonstrate that LipC is a novel cell surface-associated esterase of
M. tuberculosis
that is highly immunogenic and elicits both antibodies and cytokines/chemokines.
Association and Evidence for Linked Recognition of Type IV Secretion System Proteins VirB9-1, VirB9-2, and VirB10 in Anaplasma marginale
Authors: Morse Kaitlyn , Norimine Junzo , Palmer Guy H. , Sutten Eric L. , Baszler Timothy V. , Brown Wendy C.
Published: October 2011

Like several other bacterial pathogens,Anaplasma marginalehas an outer membrane that induces complete protection from infection and disease. However, the proteins that confer protective immunity and whether protection requires interacting proteins and/or linked T-cell and immunoglobulin G epitopes are not known. Our goal is to target the conserved type IV secretion system (T4SS) to identify conserved, immunogenic membrane proteins that are interacting and linked recognition candidates. Linked recognition is a process by which a B cell is optimally activated by a helper T cell that responds to the same, or physically associated, antigen.A. marginaleT4SS proteins VirB2, VirB4-1, VirB4-2, VirB6-1, VirB7, VirB8-2, VirB9-1, VirB9-2, VirB10, VirB11, and VirD4 were screened for their ability to induce IgG and to stimulate CD4+T cells from outer membrane-vaccinated cattle. VirB9-1, VirB9-2, and VirB10 induced the strongest IgG and T-cell responses in the majority of cattle, although three animals with major histocompatibility complex class II DRB3 restriction fragment length polymorphism types 8/23, 3/16, and 16/27 lacked T-cell responses to VirB9-1, VirB9-1 and VirB9-2, or VirB9-2 and VirB10, respectively. For these animals, VirB9-1-, VirB9-2-, and VirB10-specific IgG production may be associated with T-cell help provided by responses to an interacting protein partner(s). Interacting protein partners indicated by far-Western blotting were confirmed by immunoprecipitation assays and revealed, for the first time, specific interactions of VirB9-1 with VirB9-2 and VirB10. The immunogenicity and interactions of VirB9-1, VirB9-2, and VirB10 justify their testing as a linked protein vaccine againstA. marginale.
Innate Immune Responses and Modified Extracellular Matrix Regulation Characterize Bacterial Infection and Cellular/Connective Tissue Changes in Scarring Trachoma
Authors: Hu Victor H. , Weiss Helen A. , Ramadhani Athumani M. , Tolbert Sonda B. , Massae Patrick , Mabey David C. W. , Holland Martin J. , Bailey Robin L. , Burton Matthew J.
Published: October 2011

Trachoma is the most common infectious cause of blindness and a major public health problem in many developing countries. It is caused by recurrent ocular infection with
Chlamydia trachomatis
in childhood, with conjunctival scarring seen later in life. The pathogenesis of trachomatous scarring, however, is poorly understood, and this study was carried out to investigate the immunofibrogenic correlates of trachomatous conjunctival scarring. A case-control study of 363 cases with conjunctival scarring and 363 control participants was conducted. Investigations included
in vivo
confocal microscopy (IVCM) assessment, quantitative real-time PCR gene expression,
C. trachomatis
detection, and nonchlamydial bacterial culture. Trachomatous scarring was found to be strongly associated with a proinflammatory, innate immune response with increased expression of psoriasin, interleukin-1β, tumor necrosis factor alpha, defensin-β4A, chemokine ligand 5, and serum amyloid A1. There was also differential expression of various modifiers of the extracellular matrix, including metalloproteinases 7, 9, 10, and 12, tissue inhibitor of matrix metalloproteinase 1, and secreted protein acidic cystein-rich-like 1. The expression of many of these genes was also significantly associated with the presence of nonchlamydial bacterial infection. These infections had a marked effect on conjunctival immune processes, including an increased inflammatory infiltrate and edema seen with IVCM. This study supports the possibility that the immunofibrogenic response in scarring trachoma is partly stimulated by nonchlamydial bacterial infection, which is characterized by the expression of innate factors.
Substitutions in the BamA β-Barrel Domain Overcome the Conditional Lethal Phenotype of a Δ
bamB
Δ
bamE
Strain of Escherichia coli
Authors: Tellez Rene , Misra Rajeev
Published: October 2011

BamA interacts with the BamBCDE lipoproteins, and together they constitute the essential β-barrel assembly machine (BAM) of
Escherichia coli
. The simultaneous absence of BamB and BamE confers a conditional lethal phenotype and a severe β-barrel outer membrane protein (OMP) biogenesis defect. Without BamB and BamE, wild-type BamA levels are significantly reduced, and the folding of the BamA β-barrel, as assessed by the heat-modifiability assay, is drastically compromised. Single-amino-acid substitutions in the β-barrel domain of BamA improve both bacterial growth and OMP biogenesis in a
bamB bamE
mutant and restore BamA levels close to the BamB
+
BamE
+
level. The substitutions alter BamA β-barrel folding, and folding in the mutants becomes independent of BamB and BamE. Remarkably, BamA β-barrel alterations also improve OMP biogenesis in cells lacking the major periplasmic chaperone, SurA, which, together with BamB, is thought to facilitate the transfer of partially folded OMPs to the soluble POTRA (polypeptide-transport-associated) domain of BamA. Unlike the
bamB bamE
mutant background, the absence of BamB or SurA does not affect BamA β-barrel folding. Thus, substitutions in the outer membrane-embedded BamA β-barrel domain overcome OMP biogenesis defects that occur at the POTRA domain of BamA in the periplasm. Based on the structure of FhaC, the altered BamA residues are predicted to lie on a highly conserved loop that folds inside the β-barrel and in regions pointing outside the β-barrel, suggesting that they influence BamA function by both direct and indirect mechanisms.
Comprehensive Transcriptome Analysis of the Periodontopathogenic Bacterium Porphyromonas gingivalis W83
Authors: Høvik Hedda , Yu Wen-Han , Olsen Ingar , Chen Tsute
Published: October 2011

High-density tiling microarray and RNA sequencing technologies were used to analyze the transcriptome of the periodontopathogenic bacterium
Porphyromonas gingivalis
. The compiled
P. gingivalis
transcriptome profiles were based on total RNA samples isolated from three different laboratory culturing conditions, and the strand-specific transcription profiles generated covered the entire genome, including both protein coding and noncoding regions. The transcription profiles revealed various operon structures, 5′- and 3′-end untranslated regions (UTRs), differential expression patterns, and many novel, not-yet-annotated transcripts within intergenic and antisense regions. Further transcriptome analysis identified the majority of the genes as being expressed within operons and most 5′ and 3′ ends to be protruding UTRs, of which several 3′ UTRs were extended to overlap genes carried on the opposite/antisense strand. Extensive antisense RNAs were detected opposite most insertion sequence (IS) elements. Pairwise comparative analyses were also performed among transcriptome profiles of the three culture conditions, and differentially expressed genes and metabolic pathways were identified. With the growing realization that noncoding RNAs play important biological functions, the discovery of novel RNAs and the comprehensive transcriptome profiles compiled in this study may provide a foundation to further understand the gene regulation and virulence mechanisms in
P. gingivalis
. The transcriptome profiles can be viewed at and downloaded from the Microbial Transcriptome Database website,
http://bioinformatics.forsyth.org/mtd
.
Sudden cardiac death: mandatory exclusion of athletes at risk is a step too far
Authors: Anderson Lynley , Exeter Dan , Bowyer Lynne
Published: October 2011

Sudden cardiac death (SCD) in young athletes is a distressing event and it is not surprising that some physicians working with sports people are proposing that preventive action should be taken. There is a push for a system similar to that established in some countries, which involves screening and mandatory exclusion of those at risk. We argue that while screening can provide useful information to at-risk athletes making decisions about their future athletic careers, mandatory exclusion of athletes is paternalistic and such decisions are not rightfully within the domain of medicine.
Airway management in unconscious non-trauma patients
Authors: Nielsen Klaus , Hansen Christian Muff , Rasmussen Lars Simon
Published: October 2011

Background

Tracheal intubation is recommended in unconscious trauma patients to protect the airway from pulmonary aspiration of gastric contents and also to ensure ventilation and oxygenation. Unconsciousness is often defined as a Glasgow Coma Scale (GCS) score below 9. In non-trauma patients, however, there are no such firm recommendations regarding airway management and the GCS score may be less useful. The aim of this study was to describe the authors' experience with airway management in unconscious non-trauma patients in the prehospital setting with a physician-manned Mobile Emergency Care Unit (MECU). The main focus of the study was on the need for subsequent tracheal intubation during hospitalisation after initial treatment.

Methods

The study was based on an analysis of data prospectively collected from the MECU database in Copenhagen, Denmark. All unconscious (GCS scores below 9) non-trauma patients registered in the database during 2006 were included. The ambulance patient charts and medical records were scrutinised to assess outcome and the need for tracheal intubation during the first 24 h after admittance into hospital.

Results

A total of 557 unconscious non-trauma patients were examined and 129 patients (23%) were tracheally intubated by the MECU physician before or during transport to the hospital. Intubation was done in most patients with cardiac arrest, severe stroke or respiratory failure. Of the remaining 428 patients, 364 (85%) regained consciousness before being transported to the hospital, whereas 64 patients remained unconscious during transport and 12 (19%) of these were intubated in the emergency department.

Conclusions

The majority of unconscious non-trauma patients were not intubated in the prehospital setting. Unconscious non-trauma patients may not all need tracheal intubation before being transferred to hospital.
Best practice in primary care pathology: review 13
Authors: Cabrera-Abreu J C , Smellie W S A , Bowley R , Shaw N
Published: October 2011

This 13th best practice review examines tumour marker requesting primary care situations. The review is presented in question–answer format, referenced for each question. This review considers carcinoembryonic antigen carbohydrate antigen 15-3 (Ca15-3) and carbohydrate antigen 19-9 (Ca19-9). The recommendations represent a précis of guidance found using a standardised literature search of national and international guidance notes, consensus statements, health policy documents and evidence-based medicine reviews, supplemented by MEDLINE EMBASE searches to identify relevant primary research documents. They will be updated periodically to take account of new information.
The limits of narrative: provocations for the medical humanities
Authors: Woods Angela
Published: October 2011

This paper aims to (re)ignite debate about the role of narrative in the medical humanities. It begins with a critical review of the ways in which narrative has been mobilised by humanities and social science scholars to understand the experience of health and illness. I highlight seven dangers or blind spots in the dominant medical humanities approach to narrative, including the frequently unexamined assumption that all human beings are ‘naturally narrative’. I then explore this assumption further through an analysis of philosopher Galen Strawson's influential article ‘Against Narrativity’. Strawson rejects the descriptive claim that “human beings typically see or live or experience their lives as a narrative” and the normative claim that “a richly Narrative outlook is essential to a well-lived life, to true or full personhood”. His work has been taken up across a range of disciplines, but its implications in the context of health and illness have not yet been sufficiently discussed. This article argues that ‘Against Narrativity’ can and should stimulate robust debate within the medical humanities regarding the limits of narrative, and concludes by discussing a range of possibilities for venturing ‘beyond narrative’.
Patient based computational fluid dynamic characterization of carotid bifurcation stenosis before and after endovascular revascularization
Authors: Schirmer Clemens M , Malek Adel M
Published: October 2011

Introduction

Hemodynamic forces play a critical role in determining the molecular phenotype of the endothelial cell and in influencing vascular remodeling. A lesion based computational fluid dynamic (CFD) modeling approach is presented to understand the complex spatial and temporal hemodynamic changes that prevail in carotid stenosis (CS) in patients with critical CS undergoing carotid artery stenting (CAS).

Methods

High resolution three-dimensional rotational angiography volumetric datasets were acquired before and after treatment in eight patients, segmented and used to generate a high quality structured hexahedral computational mesh with boundary layer refinement. CFD analysis was carried out using a time dependent laminar flow model implementing non-Newtonian realistic blood viscosity for blood, and used to compute wall shear stress (WSS) and its gradient (WSSG).

Results

CAS restored fully or near laminar flow in all cases in our series. In addition, WSS was found to decrease in the stented region in all cases, reduced to near normal levels of 34±14 dyn/cm2with significant blunting of the extreme pretreatment WSSG to levels lower than 1000 dyn/cm3.

Conclusions

In this series of patients with symptomatic CS, CFD simulation enabled estimation of the hemodynamic effect of CAS, leading to reversal of abnormal flow patterns and wall shear forces around the arterial stenosis, with normalization of flow laminarity and wall shear spatiotempotal patterns known to be associated with adverse endothelial cell function.
Sensitisation to common allergens and respiratory symptoms in endotoxin exposed workers: a pooled analysis
Authors: Basinas Ioannis , Schlünssen Vivi , Heederik Dick , Sigsgaard Torben , Smit Lidwien A M , Samadi Sadegh , Omland Øyvind , Hjort Charlotte , Madsen Anne Mette , Skov Simon , Wouters Inge M
Published: October 2011

Objective

To test the hypotheses that current endotoxin exposure is inversely associated with allergic sensitisation and positively associated with non-allergic respiratory diseases in four occupationally exposed populations using a standardised analytical approach.

Methods

Data were pooled from four epidemiological studies including 3883 Dutch and Danish employees in veterinary medicine, agriculture and power plants using biofuel. Endotoxin exposure was estimated by quantitative job-exposure matrices specific for the study populations. Dose–response relationships between exposure, IgE-mediated sensitisation to common allergens and self-reported health symptoms were assessed using logistic regression and generalised additive modelling. Adjustments were made for study, age, sex, atopic predisposition, smoking habit and farm childhood. Heterogeneity was assessed by analysis stratified by study.

Results

Current endotoxin exposure was dose-dependently associated with a reduced prevalence of allergic sensitisation (ORs of 0.92, 0.81 and 0.66 for low mediate, high mediate and high exposure) and hay fever (ORs of 1.16, 0.81 and 0.58). Endotoxin exposure was a risk factor for organic dust toxic syndrome, and levels above 100 EU/m3significantly increased the risk of chronic bronchitis (p<0.0001). Stratification by farm childhood showed no effect modification except for allergic sensitisation. Only among workers without a farm childhood, endotoxin exposure was inversely associated with allergic sensitisation. Heterogeneity was primarily present for biofuel workers.

Conclusions

Occupational endotoxin exposure has a protective effect on allergic sensitisation and hay fever but increases the risk for organic dust toxic syndrome and chronic bronchitis. Endotoxin's protective effects are most clearly observed among agricultural workers.
Cancer mortality among female and male workers occupationally exposed to inorganic lead in the printing industry
Authors: Ilychova Svetlana A , Zaridze David G
Published: October 2011

Objective

Evaluation of the carcinogenicity of lead for humans has been based primarily on the results of studies on occupationally exposed men, although gender differences in lead metabolism have been reported. In addition, most of the previous studies have been limited by a failure to identify and control for co-exposures to other known occupational carcinogens. The present study follows an industrial cohort of workers, mostly women, with moderate lead exposure and no confounding by other occupational exposures.

Methods

Workers, employed at least 2 years between 1950 and 1978 in manual and mechanical (linotype) typesetting and type foundries in 27 printing plants in Moscow, were included in the cohort, which comprised 1423 men and 3102 women. The cohort was followed up during 1979–2003 and contributed 93 682 person-years of observation. Follow-up was 97.7% complete. Standardised mortality ratios (SMRs) and 95% CIs, based on mortality rates of the Moscow general population and adjusted for gender, age and calendar time, were calculated for the total cohort as well as subcohorts stratified by various exposure parameters.

Results

Among women, mortality from all causes, circulatory diseases and all cancers combined was lower than that in the Moscow general population and was similar across work groups. Among men, there was excess overall mortality, mainly due to increased mortality from ischaemic heart disease. For both sexes, no significant excess risk for any cancer site was observed, although some dose–response patterns were found. In the overall cohort, mortality from cancers of the kidney and pancreas increased up to twofold in the highest tertile of cumulative lead exposure based on duration and a relative ranking of the three subcohorts (9 deaths; SMR=2.12, 95% CI 1.10 to 4.07) and (18 deaths; SMR=2.32, 95% CI 1.46 to 3.68), respectively. Similar mortality trends for these two cancers were found in analyses by gender.

Conclusions

Consistencies by sex and exposure level make a strong case for a link between exposure to inorganic lead and cancers of the kidney and pancreas.