bluuebunny/crossref_metadata_2025_split huggingface dataset:
There and Back Again: Loss and Reacquisition of Two‐Cys Photocycles in Cyanobacteriochromes
Authors: Rockwell Nathan C. , Martin Shelley S. , Lagarias John Clark 
Published: January 2017

AbstractCyanobacteriochromes (CBCRs) are cyanobacterial photoreceptors distantly related to phytochromes. Both families use linear tetrapyrrole (bilin) chromophores that are covalently attached to a conserved Cys residue. CBCRs are more spectrally diverse than phytochromes, with known examples detecting light from the near ultraviolet to the edge of the infrared (370‐750 nm). Detection of ultraviolet to blue light by CBCRs is mediated by a second Cys residue, which forms a covalent linkage to the bilin C10 atom. Second linkage formation is best understood in a subfamily possessing a conserved Asp‐Xaa‐Cys‐Phe (DXCF) motif. Some DXCF CBCRs can isomerize their phycocyanobilin (PCB) chromophores into phycoviolobilin (PVB), a property not reported for other lineages. Both the DXCF Cys and PVB formation have been lost during evolution of other CBCR subfamilies. Using phylogenetic analysis and characterization of recombinantly expressed CBCRs, we show that the DXCF Cys residue has also been reacquired during CBCR evolution. Guided by this knowledge, we successfully reintroduced a second cysteine into a red/green CBCR, restoring blue‐light sensing and PVB formation with two additional substitutions. Our results validate the roles of these residues in CBCR spectral tuning and thus provide new insight into the molecular basis of their spectral diversity.
Population Pharmacokinetics of Piperacillin in Nonobese, Obese, and Morbidly Obese Critically Ill Patients
Authors: Alobaid Abdulaziz S. , Wallis Steven C. , Jarrett Paul , Starr Therese , Stuart Janine , Lassig-Smith Melissa , Mejia Jenny Lisette Ordóñez , Roberts Michael S. , Roger Claire , Udy Andrew A. , Lipman Jeffrey , Roberts Jason A. 
Published: January 2017

ABSTRACT

The treatment of infections in critically ill obese and morbidly obese patients is challenging because of the combined physiological changes that result from obesity and critical illness. The aim of this study was to describe the population pharmacokinetics of piperacillin in a cohort of critically ill patients, including obese and morbidly obese patients. Critically ill patients who received piperacillin-tazobactam were classified according to their body mass index (BMI) as nonobese, obese, and morbidly obese. Plasma samples were collected, and piperacillin concentrations were determined by a validated chromatographic method. Population pharmacokinetic analysis and Monte Carlo dosing simulations were performed using Pmetrics software. Thirty-seven critically ill patients (including 12 obese patients and 12 morbidly obese patients) were enrolled. The patients' mean ± standard deviation age, weight, and BMI were 50 ± 15 years, 104 ± 35 kg, and 38.0 ± 15.0 kg/m
2
, respectively. The concentration-time data were best described by a two-compartment linear model. The mean ± SD parameter estimates for the final covariate model were a clearance of 14.0 ± 7.1 liters/h, a volume of distribution of the central compartment of 49.0 ± 19.0 liters, an intercompartmental clearance from the central compartment to the peripheral compartment of 0.9 ± 0.6 liters · h
−1
, and an intercompartmental clearance from the peripheral compartment to the central compartment of 2.3 ± 2.8 liters · h
−1
. A higher measured creatinine clearance and shorter-duration infusions were associated with a lower likelihood of achieving therapeutic piperacillin exposures in patients in all BMI categories. Piperacillin pharmacokinetics are altered in the presence of obesity and critical illness. As with nonobese patients, prolonged infusions increase the likelihood of achieving therapeutic concentrations.
Subcellular Partitioning and Intramacrophage Selectivity of Antimicrobial Compounds against Mycobacterium tuberculosis
Authors: Schump Michael D. , Fox Douglas M. , Bertozzi Carolyn R. , Riley Lee W. 
Published: January 2017

ABSTRACT

The efficacy of antimicrobial drugs against
Mycobacterium tuberculosis
, an intracellular bacterial pathogen, is generally first established by testing compounds against bacteria in axenic culture. However, inside infected macrophages, bacteria encounter an environment which differs substantially from broth culture and are subject to important host-dependent pharmacokinetic phenomena which modulate drug activity. Here, we describe how pH-dependent partitioning drives asymmetric antimicrobial drug distribution in
M. tuberculosis-
infected macrophages. Specifically, weak bases with moderate activity against
M. tuberculosis
(fluoxetine, sertraline, and dibucaine) were shown to accumulate intracellularly due to differential permeability and relative abundance of their ionized and nonionized forms. Nonprotonatable analogs of the test compounds did not show this effect. Neutralization of acidic organelles directly with ammonium chloride or indirectly with bafilomycin A1 partially abrogated the growth restriction of these drugs. Using high-performance liquid chromatography, we quantified the degree of accumulation and reversibility upon acidic compartment neutralization in macrophages and observed that accumulation was greater in infected than in uninfected macrophages. We further demonstrate that the efficacy of a clinically used compound, clofazimine, is augmented by pH-based partitioning in a macrophage infection model. Because the parameters which govern this effect are well understood and are amenable to chemical modification, this knowledge may enable the rational development of more effective antibiotics against tuberculosis.
Pseudomonas aeruginosa Oligoribonuclease Contributes to Tolerance to Ciprofloxacin by Regulating Pyocin Biosynthesis
Authors: Chen Fei , Chen Gukui , Liu Yiwei , Jin Yongxin , Cheng Zhihui , Liu Yang , Yang Liang , Jin Shouguang , Wu Weihui 
Published: January 2017

ABSTRACT

Bacterial oligoribonuclease (Orn) is a conserved 3′-to-5′ exonuclease. In
Pseudomonas aeruginosa
, it has been demonstrated that Orn plays a major role in the hydrolysis of pGpG, which is required for cyclic-di-GMP homeostasis. Meanwhile, Orn is involved in the degradation of nanoRNAs, which can alter global gene expression by serving as transcription initiation primers. Previously, we found that Orn is required for the type III secretion system and pathogenesis of
P. aeruginosa
, indicating a role of Orn in the bacterial response to environmental stimuli. Here we report that Orn is required for the tolerance of
P. aeruginosa
to ciprofloxacin. Transcriptome analysis of an
orn
mutant revealed the upregulation of pyocin biosynthesis genes. Mutation of genes involved in pyocin biosynthesis in the background of an
orn
mutant restored bacterial tolerance to ciprofloxacin. We further demonstrate that the upregulation of pyocin biosynthesis genes is due to RecA-mediated autoproteolysis of PrtR, which is the major negative regulator of pyocin biosynthesis genes. In addition, the SOS response genes were upregulated in the
orn
mutant, indicating a DNA damage stress. Therefore, our results revealed a novel role of Orn in bacterial tolerance to ciprofloxacin.
Population Pharmacokinetic Model of Doxycycline Plasma Concentrations Using Pooled Study Data
Authors: Hopkins Ashley M. , Wojciechowski Jessica , Abuhelwa Ahmad Y. , Mudge Stuart , Upton Richard N. , Foster David J. R. 
Published: January 2017

ABSTRACT

The literature presently lacks a population pharmacokinetic analysis of doxycycline. This study aimed to develop a population pharmacokinetic model of doxycycline plasma concentrations that could be used to assess the power of bioequivalence between Doryx delayed-release tablets and Doryx MPC. Doxycycline pharmacokinetic data were available from eight phase 1 clinical trials following single/multiple doses of conventional-release doxycycline capsules, Doryx delayed-release tablets, and Doryx MPC under fed and fasted conditions. A population pharmacokinetic model was developed in a stepwise manner using NONMEM, version 7.3. The final covariate model was developed according to a forward inclusion (
P
< 0.01) and then backward deletion (
P
< 0.001) procedure. The final model was a two-compartment model with two-transit absorption compartments. Structural covariates in the base model included formulation effects on relative bioavailability (
F
), absorption lag (ALAG), and the transit absorption rate (KTR) under the fed status. An absorption delay (lag) for the fed status (FTLAG2 = 0.203 h) was also included in the model as a structural covariate. The fed status was observed to decrease
F
by 10.5%, and the effect of female sex was a 14.4% increase in clearance. The manuscript presents the first population pharmacokinetic model of doxycycline plasma concentrations following oral doxycycline administration. The model was used to assess the power of bioequivalence between Doryx delayed-release tablets and Doryx MPC, and it could potentially be used to critically examine and optimize doxycycline dose regimens.
Local Delivery of the Toll-Like Receptor 9 Ligand CpG Downregulates Host Immune and Inflammatory Responses, Ameliorating Established Leishmania (Viannia) panamensis Chronic Infection
Authors: Ehrlich Allison K. , Fernández Olga L. , Rodriguez-Pinto Daniel , Castilho Tiago M. , Corral Caridad Maria J. , Goldsmith-Pestana Karen , Saravia Nancy Gore , McMahon-Pratt Diane 
Published: January 2017

ABSTRACTInfection byLeishmania(Viannia)panamensis, the predominant etiologic agent for cutaneous leishmaniasis in Colombia, is characterized by a chronic mixed inflammatory response. Current treatment options are plagued by toxicity, lengthy treatment regimens, and growing evidence of drug resistance. Immunotherapy, modulating the immune system to mount a protective response, may provide an alternate therapeutic approach. We investigated the ability of the Toll-like receptor 9 (TLR9) ligand CpG to modulate established disease in theL. (V.)panamensismouse model. Treatment of established infection with a high dose (50 μg) of CpG ameliorated disease and lowered parasite burden. Interestingly, immediately after treatment there was a significant increase in transforming growth factor β (TGF-β) and concomitantly an increase in T regulatory cell (Treg) function. Although a general reduction in cell-mediated immune cytokine and chemokine (gamma interferon [IFN-γ], interleukin 10 [IL-10], IL-13, IL-6, granulocyte-macrophage colony-stimulating factor [GM-CSF], IL-4, and MIP-1α) responses of the treated mice was observed, certain chemokines (RANTES, monocyte chemoattractant protein 1[MCP-1], and IP-10) were increased. Further, in peripheral blood mononuclear cells (PBMCs) from patients with cutaneous leishmaniasis, CpG treatment similarly exhibited a dose-response effect on the production of IFN-γ, IL-17, IL-10, and IL-13, with reductions observed at higher doses. To further understand the underlying mechanisms and cell populations driving the CpG mediated response, we examined theex vivodose effects mediated by the TLR9+cell populations (dendritic cells, macrophages, and B cells) found to accumulate labeled CpGin vivo. Notably, B cells altered the production of IL-17, IL-13, and IFN-γ, supporting a role for B cells functioning as antigen-presenting cells (APCs) and/or regulatory cells during infection. Interestingly, B cells have been previously demonstrated as a primary type of APC in patients infected withL. (V.)panamensisand thus may be useful targets of immunotherapy. Collectively, our results show that CpG-induced immune regulation leads to a dampening of the host immune response and healing in the mouse model, and it may provide an alternate approach to treatment of cutaneous leishmaniasis caused byL. (V.)panamensis.
Characterization and Diagnostic Application of Trypanosoma cruzi Trypomastigote Excreted-Secreted Antigens Shed in Extracellular Vesicles Released from Infected Mammalian Cells
Authors: Bautista-López Norma L. , Ndao Momar , Camargo Fabio Vasquez , Nara Takeshi , Annoura Takeshi , Hardie Darryl B. , Borchers Christoph H. , Jardim Armando 
Published: January 2017

ABSTRACTChagas disease, caused byTrypanosoma cruzi, although endemic in many parts of Central and South America, is emerging as a global health threat through the potential contamination of blood supplies. Consequently, in the absence of a gold standard assay for the diagnosis of Chagas disease, additional antigens or strategies are needed. A proteomic analysis of the trypomastigote excreted-secreted antigens (TESA) associated with exosomal vesicles shed byT. cruziidentified ∼80 parasite proteins, with the majority beingtrans-sialidases. Mass spectrometry analysis of immunoprecipitation products performed using Chagas immune sera showed a marked enrichment in a subset of TESA proteins. Of particular relevance for diagnostic applications were the retrotransposon hot spot (RHS) proteins, which are absent inLeishmaniaspp., parasites that often confound diagnosis of Chagas disease. Interestingly, serological screens using recombinant RHS showed a robust immunoreactivity with sera from patients with clinical stages of Chagas ranging from asymptomatic to advance cardiomyopathy and this immunoreactivity was comparable to that of crude TESA. More importantly, no cross-reactivity with RHS was detected with sera from patients with malaria, leishmaniasis, toxoplasmosis, or African sleeping sickness, making this protein an attractive reagent for diagnosis of Chagas disease.
Improved Differentiation of Streptococcus pneumoniae and Other S. mitis Group Streptococci by MALDI Biotyper Using an Improved MALDI Biotyper Database Content and a Novel Result Interpretation Algorithm
Authors: Harju Inka , Lange Christoph , Kostrzewa Markus , Maier Thomas , Rantakokko-Jalava Kaisu , Haanperä Marjo 
Published: January 2017

ABSTRACTReliable distinction ofStreptococcus pneumoniaeand viridans group streptococci is important because of the different pathogenic properties of these organisms. Differentiation betweenS. pneumoniaeand closely relatedSreptococcusmitisspecies group streptococci has always been challenging, even when using such modern methods as 16S rRNA gene sequencing or matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry. In this study, a novel algorithm combined with an enhanced database was evaluated for differentiation betweenS. pneumoniaeandS. mitisspecies group streptococci. One hundred one clinicalS. mitisspecies group streptococcal strains and 188 clinicalS. pneumoniaestrains were identified by both the standard MALDI Biotyper database alone and that combined with a novel algorithm. The database update from 4,613 strains to 5,627 strains drastically improved the differentiation ofS. pneumoniaeandS. mitisspecies group streptococci: when the new database version containing 5,627 strains was used, only one of the 101S. mitisspecies group isolates was misidentified asS. pneumoniae, whereas 66 of them were misidentified asS. pneumoniaewhen the earlier 4,613-strain MALDI Biotyper database version was used. The updated MALDI Biotyper database combined with the novel algorithm showed even better performance, producing no misidentifications of theS. mitisspecies group strains asS. pneumoniae. AllS. pneumoniaestrains were correctly identified asS. pneumoniaewith both the standard MALDI Biotyper database and the standard MALDI Biotyper database combined with the novel algorithm. This new algorithm thus enables reliable differentiation between pneumococci and otherS. mitisspecies group streptococci with the MALDI Biotyper.
The Bactec FX Blood Culture System Detects Brucella melitensis Bacteremia in Adult Patients within the Routine 1-Week Incubation Period
Authors: Sagi Moshe , Nesher Lior , Yagupsky Pablo 
Published: January 2017

ABSTRACT

The performance of the Bactec FX blood culture system for detecting
Brucella
bacteremia within the routine 1-week incubation period was assessed in a prospective study conducted in an area in southern Israel in which
Brucella melitensis
is endemic. Aerobic vials (BD Bactec Plus Aerobic/F medium) inoculated with blood specimens obtained from adult patients with positive Rose-Bengal screening test results were monitored for 4 consecutive weeks, and blind subcultures of negative vials were performed on solid media on days 7 and 28. During a 16-month period, a total of 31 (35.2%) of 88 cultures, obtained from 19 (38.0%) of 50 patients, were positive for
Brucella melitensis
. The blood culture instrument identified 30 (96.8%) of 31 positive vials within 7 days of incubation; the single positive vial that was missed by the automated readings was detected only by the blind subculture performed on day 28. It is concluded that the Bactec FX system is able to detect the vast majority of episodes of
Brucella
bacteremia within the 1-week incubation protocol instituted in most clinical microbiology laboratories and without the need to perform blind subcultures of negative vials, enabling early diagnosis and saving labor and incubation time and space.
Human Cytomegalovirus (HCMV)-Specific CD4
+
T Cells Are Polyfunctional and Can Respond to HCMV-Infected Dendritic Cells
In Vitro
Authors: Jackson Sarah E. , Sedikides George X. , Mason Gavin M. , Okecha Georgina , Wills Mark R. 
Published: January 2017

ABSTRACT

Human cytomegalovirus (HCMV) infection and periodic reactivation are generally well controlled by the HCMV-specific T cell response in healthy people. While the CD8
+
T cell response to HCMV has been extensively studied, the HCMV-specific CD4
+
T cell effector response is not as well understood, especially in the context of direct interactions with HCMV-infected cells. We screened the gamma interferon (IFN-γ) and interleukin-10 (IL-10) responses to 6 HCMV peptide pools (pp65, pp71, IE1, IE2, gB, and US3, selected because they were the peptides most frequently responded to in our previous studies) in 84 donors aged 23 to 74 years. The HCMV-specific CD4
+
T cell response to pp65, IE1, IE2, and gB was predominantly Th1 biased, with neither the loss nor the accumulation of these responses occurring with increasing age. A larger proportion of donors produced an IL-10 response to pp71 and US3, but the IFN-γ response was still dominant. CD4
+
T cells specific to the HCMV proteins studied were predominantly effector memory cells and produced both cytotoxic (CD107a expression) and cytokine (macrophage inflammatory protein 1β secretion) effector responses. Importantly, when we measured the CD4
+
T cell response to cytomegalovirus (CMV)-infected dendritic cells
in vitro
, we observed that the CD4
+
T cells produced a range of cytotoxic and secretory effector functions, despite the presence of CMV-encoded immune evasion molecules. CD4
+
T cell responses to HCMV-infected dendritic cells were sufficient to control the dissemination of virus in an
in vitro
assay. Together, the results show that HCMV-specific CD4
+
T cell responses, even those from elderly individuals, are highly functional and are directly antiviral.

IMPORTANCE
Human cytomegalovirus (HCMV) infection is carried for a lifetime and in healthy people is kept under control by the immune system. HCMV has evolved many mechanisms to evade the immune response, possibly explaining why the virus is never eliminated during the host's lifetime. The dysfunction of immune cells associated with the long-term carriage of HCMV has been linked with poor responses to new pathogens and vaccines when people are older. In this study, we investigated the response of a subset of immune cells (CD4
+
T cells) to HCMV proteins in healthy donors of all ages, and we demonstrate that the functionality of CD4
+
T cells is maintained. We also show that CD4
+
T cells produce effector functions in response to HCMV-infected cells and can prevent virus spread. Our work demonstrates that these HCMV-specific immune cells retain many important functions and help to prevent deleterious HCMV disease in healthy older people.
Subcellular Localization of HIV-1
gag-pol
mRNAs Regulates Sites of Virion Assembly
Authors: Becker Jordan T. , Sherer Nathan M. 
Published: January 2017

ABSTRACT

Full-length unspliced human immunodeficiency virus type 1 (HIV-1) RNAs serve dual roles in the cytoplasm as mRNAs encoding the Gag and Gag-Pol capsid proteins as well as genomic RNAs (gRNAs) packaged by Gag into virions undergoing assembly at the plasma membrane (PM). Because Gag is sufficient to drive the assembly of virus-like particles even in the absence of gRNA binding, whether viral RNA trafficking plays an active role in the native assembly pathway is unknown. In this study, we tested the effects of modulating the cytoplasmic abundance or distribution of full-length viral RNAs on Gag trafficking and assembly in the context of single cells. Increasing full-length viral RNA abundance or distribution had little-to-no net effect on Gag assembly competency when provided in
trans
. In contrast, artificially tethering full-length viral RNAs or surrogate
gag-pol
mRNAs competent for Gag synthesis to non-PM membranes or the actin cytoskeleton severely reduced net virus particle production. These effects were explained, in large part, by RNA-directed changes to Gag's distribution in the cytoplasm, yielding aberrant subcellular sites of virion assembly. Interestingly, RNA-dependent disruption of Gag trafficking required either of two
cis
-acting RNA regulatory elements: the 5′ packaging signal (Psi) bound by Gag during genome encapsidation or, unexpectedly, the Rev response element (RRE), which regulates the nuclear export of gRNAs and other intron-retaining viral RNAs. Taken together, these data support a model for native infection wherein structural features of the
gag-pol
mRNA actively compartmentalize Gag to preferred sites within the cytoplasm and/or PM.

IMPORTANCE
The spatial distribution of viral mRNAs within the cytoplasm can be a crucial determinant of efficient translation and successful virion production. Here we provide direct evidence that mRNA subcellular trafficking plays an important role in regulating the assembly of human immunodeficiency virus type 1 (HIV-1) virus particles at the plasma membrane (PM). Artificially tethering viral mRNAs encoding Gag capsid proteins (
gag-pol
mRNAs) to distinct non-PM subcellular locales, such as cytoplasmic vesicles or the actin cytoskeleton, markedly alters Gag subcellular distribution, relocates sites of assembly, and reduces net virus particle production. These observations support a model for native HIV-1 assembly wherein HIV-1
gag-pol
mRNA localization helps to confine interactions between Gag, viral RNAs, and host determinants in order to ensure virion production at the right place and right time. Direct perturbation of HIV-1 mRNA subcellular localization may represent a novel antiviral strategy.