bluuebunny/crossref_metadata_2025_split huggingface dataset:
Structure-Guided Expansion of the Substrate Range of Methylmalonyl Coenzyme A Synthetase (MatB) of Rhodopseudomonas palustris
Authors: Crosby Heidi A. , Rank Katherine C. , Rayment Ivan , Escalante-Semerena Jorge C. 
Published: July 2012

ABSTRACTMalonyl coenzyme A (malonyl-CoA) and methylmalonyl-CoA are two of the most commonly used extender units for polyketide biosynthesis and are utilized to synthesize a vast array of pharmaceutically relevant products with antibacterial, antiparasitic, anticholesterol, anticancer, antifungal, and immunosuppressive properties. Heterologous hosts used for polyketide production such asEscherichia colioften do not produce significant amounts of methylmalonyl-CoA, however, requiring the introduction of other pathways for the generation of this important building block. Recently, the bacterial malonyl-CoA synthetase class of enzymes has been utilized to generate malonyl-CoA and methylmalonyl-CoA directly from malonate and methylmalonate. We demonstrate that in the purple photosynthetic bacteriumRhodopseudomonas palustris, MatB (RpMatB) acts as a methylmalonyl-CoA synthetase and is required for growth on methylmalonate. We report theapo(1.7-Å resolution) and ATP-bound (2.0-Å resolution) structure and kinetic analysis ofRpMatB, which shows similar activities for both malonate and methylmalonate, making it an ideal enzyme for heterologous polyketide biosynthesis. Additionally, rational, structure-based mutagenesis of the active site ofRpMatB led to substantially higher activity with ethylmalonate and butylmalonate, demonstrating that this enzyme is a prime target for expanded substrate specificity.
Comparative Analysis of Mycobacterial Truncated Hemoglobin Promoters and thegroEL2Promoter in Free-Living and Intracellular Mycobacteria
Authors: Joseph Sunil V. , Madhavilatha G. K. , Kumar R. Ajay , Mundayoor Sathish 
Published: July 2012

ABSTRACTThe success ofMycobacterium tuberculosisdepends on its ability to withstand and survive the hazardous environment inside the macrophages that are created by reactive oxygen intermediates, reactive nitrogen intermediates, severe hypoxia, low pH, and high CO2levels. Therefore, an effective detoxification system is required for the pathogen to persistin vivo. The genome ofM. tuberculosiscontains a new family of hemoproteins named truncated hemoglobin O (trHbO) and truncated hemoglobin N (trHbN), encoded by theglbOandglbNgenes, respectively, important in the survival ofM. tuberculosisin macrophages. Mycobacterial heat shock proteins are known to undergo rapid upregulation under stress conditions. The expression profiles of the promoters of these genes were studied by constructing transcriptional fusions with green fluorescent protein and monitoring the promoter activity in both free-living and intracellular milieus at different time points. WhereasglbNshowed an early response to the oxidative and nitrosative stresses tested,glbOgave a lasting response to lower concentrations of both stresses. At all time points and under all stress conditions tested,groEL2showed higher expression than both trHb promoters and expression of both promoters showed an increase while inside the macrophages. Real-time PCR analysis of trHb andgroEL2mRNAs showed an initial upregulation at 24 h postinfection. The presence of theglbOprotein imparted an increased survival toM. smegmatisin THP-1 differentiated macrophages compared to that imparted by theglbNandhsp65proteins. The comparative upregulation shown by both trHb promoters while grown inside macrophages indicates the importance of these promoters for the survival ofM. tuberculosisin the hostile environment of the host.
Pediatric Invasive Pneumococcal Disease in the United States in the Era of Pneumococcal Conjugate Vaccines
Authors: Tan Tina Q. 
Published: July 2012

SUMMARY

Invasive infections caused by
Streptococcus pneumoniae
continue to be a major cause of morbidity and mortality worldwide, especially in children under 5 years of age. In the United States, 90% of invasive pneumococcal infections in children are caused by 13 serotypes of
S. pneumoniae
. The licensure (in 2000) and subsequent widespread use of a heptavalent pneumococcal conjugate vaccine (PCV7) have had a significant impact on decreasing the incidence of serious invasive pneumococcal disease (IPD) in all age groups, especially in children under 2 years of age. However, the emergence of replacement non-PCV7 serotypes, especially serotype 19A, has resulted in an increase in the incidence of serious and invasive infections. In 2010, a 13-valent PCV was licensed in the United States. However, the impact that this vaccine will have on IPD remains to be seen. The objectives of this review are to discuss the epidemiology of serious and invasive pneumococcal infections in the United States in the PCV era and to review some of the pneumococcal vaccines that are in development.
Antimicrobial Susceptibility Testing, Drug Resistance Mechanisms, and Therapy of Infections with Nontuberculous Mycobacteria
Authors: Brown-Elliott Barbara A. , Nash Kevin A. , Wallace Richard J. 
Published: July 2012

SUMMARY
Within the past 10 years, treatment and diagnostic guidelines for nontuberculous mycobacteria have been recommended by the American Thoracic Society (ATS) and the Infectious Diseases Society of America (IDSA). Moreover, the Clinical and Laboratory Standards Institute (CLSI) has published and recently (in 2011) updated recommendations including suggested antimicrobial and susceptibility breakpoints. The CLSI has also recommended the broth microdilution method as the gold standard for laboratories performing antimicrobial susceptibility testing of nontuberculous mycobacteria. This article reviews the laboratory, diagnostic, and treatment guidelines together with established and probable drug resistance mechanisms of the nontuberculous mycobacteria.
Enteric Protozoa in the Developed World: a Public Health Perspective
Authors: Fletcher Stephanie M. , Stark Damien , Harkness John , Ellis John 
Published: July 2012

SUMMARY
Several enteric protozoa cause severe morbidity and mortality in both humans and animals worldwide. In developed settings, enteric protozoa are often ignored as a cause of diarrheal illness due to better hygiene conditions, and as such, very little effort is used toward laboratory diagnosis. Although these protozoa contribute to the high burden of infectious diseases, estimates of their true prevalence are sometimes affected by the lack of sensitive diagnostic techniques to detect them in clinical and environmental specimens. Despite recent advances in the epidemiology, molecular biology, and treatment of protozoan illnesses, gaps in knowledge still exist, requiring further research. There is evidence that climate-related changes will contribute to their burden due to displacement of ecosystems and human and animal populations, increases in atmospheric temperature, flooding and other environmental conditions suitable for transmission, and the need for the reuse of alternative water sources to meet growing population needs. This review discusses the common enteric protozoa from a public health perspective, highlighting their epidemiology, modes of transmission, prevention, and control. It also discusses the potential impact of climate changes on their epidemiology and the issues surrounding waterborne transmission and suggests a multidisciplinary approach to their prevention and control.
Sensitivity and Specificity of an Operon Immunochromatographic Test in Serum and Whole-Blood Samples for the Diagnosis of Trypanosoma cruzi Infection in Spain, an Area of Nonendemicity
Authors: Flores-Chavez María , Cruz Israel , Nieto Javier , Gárate Teresa , Navarro Miriam , Pérez-Ayala Ana , López-Vélez Rogelio , Cañavate Carmen 
Published: July 2012

ABSTRACT

Trypanosoma cruzi
infection is an imported parasitic disease in Spain, and the majority of infected individuals are in the chronic phase of the disease. This study evaluated the sensitivity and specificity of the Operon immunochromatographic test (ICT-Operon; Simple Stick Chagas and Simple Chagas WB [whole blood]; Operon S.A., Spain) for different biological samples. Well-characterized serum samples were obtained from chagasic patients (
n
= 63), nonchagasic individuals (
n
= 95), visceral leishmaniasis patients (
n
= 38), and malaria patients (
n
= 55). Noncharacterized specimens were obtained from Latin American immigrants and individuals at risk with a clinical and/or epidemiological background: these specimens were recovered serum or plasma samples (
n
= 450), whole peripheral blood (
n
= 94), and capillary blood (
n
= 282). The concordance of the results by enzyme-linked immunosorbent assay and indirect immunofluorescence test was considered to be the “gold standard” for diagnosis. Serum and plasma samples were analyzed by Stick Chagas, and whole blood was analyzed by Simple Chagas WB. The sensitivity and specificity of the ICT-Operon in well-characterized samples were 100% and 97.9%, respectively. No cross-reactivity was found with samples obtained from visceral leishmaniasis patients. In contrast, a false-positive result was obtained in 27.3% of samples from malaria patients. The sensitivities of the rapid test in noncharacterized serum or plasma, peripheral blood, and capillary blood samples were 100%, 92.1%, and 86.4%, respectively, while the specificities were 91.6%, 93.6%, and 95% in each case. ICT-Operon showed variable sensitivity, depending on the kind of sample, performing better when serum or plasma samples were used. It could therefore be used for serological screening combined with any other conventional test.
Goats Primed with Mycobacterium bovis BCG and Boosted with a Recombinant Adenovirus Expressing Ag85A Show Enhanced Protection against Tuberculosis
Authors: Pérez de Val Bernat , Villarreal-Ramos Bernardo , Nofrarías Miquel , López-Soria Sergio , Romera Nadine , Singh Mahavir , Abad F. Xavier , Xing Zhou , Vordermeier H. Martin , Domingo Mariano 
Published: July 2012

ABSTRACTThis is the first efficacy study using the experimental goat model, a natural host of tuberculosis (TB), to evaluate the efficacy of heterologousMycobacterium bovisbacillus Calmette-Guérin (BCG) prime followed by boosting with a replication-deficient adenovirus expressing the antigen Ag85A (AdAg85A). Three experimental groups of 11 goat kids each were used: BCG vaccinated, BCG vaccinated and AdAg85A boosted, and nonvaccinated. Twenty-two goat kids were vaccinated with ∼5 × 105CFU of BCG (week 0), and 11 of them were boosted at week 8 with 109PFU of AdAg85A. At week 14, all goats were challenged by the endobronchial route with ∼1.5 × 103CFU ofMycobacterium caprae. The animals were euthanized at week 28. Cellular and humoral immunity induced by vaccination andM. capraeinfection was measured throughout the study. After challenge BCG-AdAg85A-vaccinated animals exhibited reduced pathology compared to BCG-vaccinated animals in lungs and in pulmonary lymph nodes. There were significant reductions in bacterial load in both groups of vaccinated goats, but the reduction was more pronounced in prime-boosted animals. Antigen-specific gamma interferon (IFN-γ) and humoral responses were identified as prognostic biomarkers of vaccination outcome depending on their correlation with pathological and bacteriological results. As far as we know, this is the first report using multidetector computed tomography (MDCT) to measure vaccine efficacy against pulmonary TB in an animal model. The use in vaccine trials of animals that are natural hosts of TB may improve research into human TB vaccines.
Lipopolysaccharide Heterogeneity in the Atypical Group of Novel Emerging Brucella Species
Authors: Zygmunt Michel S. , Jacques Isabelle , Bernardet Nelly , Cloeckaert Axel 
Published: July 2012

ABSTRACT

Recently, novel
Brucella
strains with phenotypic characteristics that were atypical for strains belonging to the genus
Brucella
have been reported. Phenotypically many of these strains were initially misidentified as
Ochrobactrum
spp. Two novel species have been described so far for these strains, i.e.,
B. microti
and
B. inopinata
, and other strains genetically related to
B. inopinata
may constitute other novel species as well. In this study, we analyzed the lipopolysaccharides (LPS) (smooth LPS [S-LPS] and rough LPS [R-LPS]) of these atypical strains using different methods and a panel of monoclonal antibodies (MAbs) directed against several epitopes of the
Brucella
O-polysaccharide (O-PS) and R-LPS. Among the most striking results,
Brucella
sp. strain BO2, isolated from a patient with chronic destructive pneumonia, showed a completely distinct S-LPS profile in silver stain gels that looked more similar to that of enterobacterial S-LPS. This strain also failed to react with MAbs against
Brucella
O-PS epitopes and showed weak reactivity with anti-R-LPS MAbs.
B. inopinata
reference strain BO1 displayed an M-dominant S-LPS type with some heterogeneity relative to the classical M-dominant
Brucella
S-LPS type. Australian wild rodent strains belonging also to the
B. inopinata
group showed a classical A-dominant S-LPS but lacked the O-PS common (C) epitopes, as previously reported for
B. suis
biovar 2 strains. Interestingly, some strains also failed to react with anti-R-LPS MAbs, such as the
B. microti
reference strain and
B. inopinata
BO1, suggesting modifications in the core-lipid A moieties of these strains. These results have several implications for serological typing and serological diagnosis and underline the need for novel tools for detection and correct identification of such novel emerging
Brucella
spp.
Schizosaccharomyces pombe Hat1 (Kat1) Is Associated with Mis16 and Is Required for Telomeric Silencing
Authors: Tong Kevin , Keller Thomas , Hoffman Charles S. , Annunziato Anthony T. 
Published: July 2012

ABSTRACT

The Hat1 histone acetyltransferase has been implicated in the acetylation of histone H4 during chromatin assembly. In this study, we have characterized the Hat1 complex from the fission yeast
Schizosaccharomyces pombe
and have examined its role in telomeric silencing. Hat1 is found associated with the RbAp46 homologue Mis16, an essential protein. The Hat1 complex acetylates lysines 5 and 12 of histone H4, the sites that are acetylated in newly synthesized H4 in a wide range of eukaryotes. Deletion of
hat1
in
S. pombe
is itself sufficient to cause the loss of silencing at telomeres. This is in contrast to results obtained with an
S. cerevisiae hat1Δ
strain, which must also carry mutations of specific acetylatable lysines in the H3 tail domain for loss of telomeric silencing to occur. Notably, deletion of
hat1
from
S. pombe
resulted in an increase of acetylation of histone H4 in subtelomeric chromatin, concomitant with derepression of this region. A similar loss of telomeric silencing was also observed after growing cells in the presence of the deacetylase inhibitor trichostatin A. However, deleting
hat1
did not cause loss of silencing at centromeres or the silent mating type locus. These results point to a direct link between Hat1, H4 acetylation, and the establishment of repressed telomeric chromatin in fission yeast.
Dual RpoH Sigma Factors and Transcriptional Plasticity in a Symbiotic Bacterium
Authors: Barnett Melanie J. , Bittner Alycia N. , Toman Carol J. , Oke Valerie , Long Sharon R. 
Published: July 2012

ABSTRACTSinorhizobium melilotican live as a soil saprophyte and can engage in a nitrogen-fixing symbiosis with plant roots. To succeed in such diverse environments, the bacteria must continually adjust gene expression. Transcriptional plasticity in eubacteria is often mediated by alternative sigma (σ) factors interacting with core RNA polymerase. TheS. melilotigenome encodes 14 of these alternative σ factors, including two putative RpoH (“heat shock”) σ factors. We used custom Affymetrix symbiosis chips to characterize the global transcriptional response ofS. melilotirpoH1,rpoH2, andrpoH1 rpoH2mutants during heat shock and stationary-phase growth. Under these conditions, expression of over 300 genes is dependent onrpoH1andrpoH2. We mapped transcript start sites of 69rpoH-dependent genes using 5′ RACE (5′ rapid amplification of cDNA ends), which allowed us to determine putative RpoH1-dependent, RpoH2-dependent, and dual-promoter (RpoH1- and RpoH2-dependent) consensus sequences that were each used to search the genome for other potential direct targets of RpoH. The inferredS. melilotiRpoH promoter consensus sequences share features ofEscherichia coliRpoH promoters but lack extended −10 motifs.
A Single Amino Acid Substitution Changes the Self-Assembly Status of a Type IV Piliation Secretin
Authors: Nickerson Nicholas N. , Abby Sophie S. , Rocha Eduardo P. C. , Chami Mohamed , Pugsley Anthony P. 
Published: July 2012

ABSTRACTSecretins form large multimeric complexes in the outer membranes of many Gram-negative bacteria, where they function as dedicated gateways that allow proteins to access the extracellular environment. Despite their overall relatedness, different secretins use different specific and general mechanisms for their targeting, assembly, and membrane insertion. We report that all tested secretins from several type II secretion systems and from the filamentous bacteriophage f1 can spontaneously multimerize and insert into liposomes in anin vitrotranscription-translation system. Phylogenetic analyses indicate that these secretins form a group distinct from the secretins of the type IV piliation and type III secretion systems, which do not autoassemblein vitro. A mutation causing a proline-to-leucine substitution allowed PilQ secretins from two different type IV piliation systems to assemblein vitro, albeit with very low efficiency, suggesting that autoassembly is an inherent property of all secretins.
Novel Listerial Glycerol Dehydrogenase- and Phosphoenolpyruvate-Dependent Dihydroxyacetone Kinase System Connected to the Pentose Phosphate Pathway
Authors: Monniot Céline , Zébré Arthur Constant , Aké Francine Moussan Désirée , Deutscher Josef , Milohanic Eliane 
Published: July 2012

ABSTRACTSeveral bacteria use glycerol dehydrogenase to transform glycerol into dihydroxyacetone (Dha). Dha is subsequently converted into Dha phosphate (Dha-P) by an ATP- or phosphoenolpyruvate (PEP)-dependent Dha kinase.Listeria innocuapossesses two potential PEP-dependent Dha kinases. One is encoded by 3 of the 11 genes forming the glycerol (gol) operon. This operon also containsgolD(lin0362), which codes for a new type of Dha-forming NAD+-dependent glycerol dehydrogenase. The subsequent metabolism of Dha requires its phosphorylation via the PEP:sugar phosphotransferase system components enzyme I, HPr, and EIIADha-2 (Lin0369). P∼EIIADha-2 transfers its phosphoryl group to DhaL-2, which phosphorylates Dha bound to DhaK-2. The resulting Dha-P is probably metabolized mainly via the pentose phosphate pathway, because two genes of thegoloperon encode proteins resembling transketolases and transaldolases. In addition, purified Lin0363 and Lin0364 exhibit ribose-5-P isomerase (RipB) and triosephosphate isomerase activities, respectively. The latter enzyme converts part of the Dha-P into glyceraldehyde-3-P, which, together with Dha-P, is metabolized via gluconeogenesis to form fructose-6-P. Together with another glyceraldehyde-3-P molecule, the transketolase transforms fructose-6-P into intermediates of the pentose phosphate pathway. Thegoloperon is preceded bygolR, transcribed in the opposite orientation and encoding a DeoR-type repressor. Its inactivation causes the constitutive but glucose-repressible expression of the entiregoloperon, including the last gene, encoding a pediocin immunity-like (PedB-like) protein. Its elevated level of synthesis in thegolRmutant causes slightly increased immunity against pediocin PA-1 compared to the wild-type strain or apedB-like deletion mutant.
Nucleotides Critical for the Interaction of the Streptococcus pyogenes Mga Virulence Regulator with Mga-Regulated Promoter Sequences
Authors: Hause Lara L. , McIver Kevin S. 
Published: July 2012

ABSTRACTThe Mga regulator ofStreptococcus pyogenesdirectly activates the transcription of a core regulon that encodes virulence factors such as M protein (emm), C5a peptidase (scpA), and streptococcal inhibitor of complement (sic) by directly binding to a 45-bp binding site as determined by an electrophoretic mobility shift assay (EMSA) and DNase I protection. However, by comparing the nucleotide sequences of all established Mga binding sites, we found that they exhibit only 13.4% identity with no discernible symmetry. To determine the core nucleotides involved in functional Mga-DNA interactions, the M1T1 Pemm1binding site was altered and screened for nucleotides important for DNA bindingin vitroand for transcriptional activation using a plasmid-based luciferase reporterin vivo. Following this analysis, 34 nucleotides within the Pemm1binding site that had an effect on Mga binding, Mga-dependent transcriptional activation, or both were identified. Of these critical nucleotides, guanines and cytosines within the major groove were disproportionately identified clustered at the 5′ and 3′ ends of the binding site and with runs of nonessential adenines between the critical nucleotides. On the basis of these results, a Pemm1minimal binding site of 35 bp bound Mga at a level comparable to the level of binding of the larger 45-bp site. Comparison of Pemmwith directed mutagenesis performed in the M1T1 Mga-regulated PscpAand Psicpromoters, as well as methylation interference analysis of PscpA, establish that Mga binds to DNA in a promoter-specific manner.
Physical Interaction between Coat Morphogenetic Proteins SpoVID and CotE Is Necessary for Spore Encasement in Bacillus subtilis
Authors: de Francesco Melissa , Jacobs Jake Z. , Nunes Filipa , Serrano Mónica , McKenney Peter T. , Chua Ming-Hsiu , Henriques Adriano O. , Eichenberger Patrick 
Published: July 2012

ABSTRACTEndospore formation byBacillus subtilisis a complex and dynamic process. One of the major challenges of sporulation is the assembly of a protective, multilayered, proteinaceous spore coat, composed of at least 70 different proteins. Spore coat formation can be divided into two distinct stages. The first is the recruitment of proteins to the spore surface, dependent on the morphogenetic protein SpoIVA. The second step, known as encasement, involves the migration of the coat proteins around the circumference of the spore in successive waves, a process dependent on the morphogenetic protein SpoVID and the transcriptional regulation of individual coat genes. We provide genetic and biochemical evidence supporting the hypothesis that SpoVID promotes encasement of the spore by establishing direct protein-protein interactions with other coat morphogenetic proteins. It was previously demonstrated that SpoVID directly interacts with SpoIVA and the inner coat morphogenetic protein, SafA. Here, we show by yeast two-hybrid and pulldown assays that SpoVID also interacts directly with the outer coat morphogenetic protein, CotE. Furthermore, by mutational analysis, we identified a specific residue in the N-terminal domain of SpoVID that is essential for the interaction with CotE but dispensable for the interaction with SafA. We propose an updated model of coat assembly and spore encasement that incorporates several physical interactions between the principal coat morphogenetic proteins.