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Emerging Infectious Diseases Sep 2017FosA, a glutathione S-transferase that inactivates fosfomycin, has been reported as the cause of enzymatic resistance to fosfomycin. We show that multiple lineages of...
FosA, a glutathione S-transferase that inactivates fosfomycin, has been reported as the cause of enzymatic resistance to fosfomycin. We show that multiple lineages of FosA-producing extended spectrum β-lactamase Escherichia coli have circulated in France since 2012, potentially reducing the efficacy of fosfomycin in treating infections with antimicrobial drug-resistant gram-negative bacilli.
Topics: Anti-Bacterial Agents; Drug Resistance, Multiple, Bacterial; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Foscarnet; Fosfomycin; France; Gene Expression; Humans; Isoenzymes; Microbial Sensitivity Tests; Plasmids; Prevalence; beta-Lactamases
PubMed: 28820368
DOI: 10.3201/eid2309.170560 -
Proceedings of the National Academy of... Aug 2017The enzyme 1-deoxy-d-xylulose 5-phosphate synthase (DXPS) is a key enzyme in the methylerythritol 4-phosphate pathway and is a target for the development of antibiotics,...
The enzyme 1-deoxy-d-xylulose 5-phosphate synthase (DXPS) is a key enzyme in the methylerythritol 4-phosphate pathway and is a target for the development of antibiotics, herbicides, and antimalarial drugs. DXPS catalyzes the formation of 1-deoxy-d-xylulose 5-phosphate (DXP), a branch point metabolite in isoprenoid biosynthesis, and is also used in the biosynthesis of thiamin (vitamin B) and pyridoxal (vitamin B). Previously, we found that DXPS is unique among the superfamily of thiamin diphosphate (ThDP)-dependent enzymes in stabilizing the predecarboxylation intermediate, C2-alpha-lactyl-thiamin diphosphate (LThDP), which has subsequent decarboxylation that is triggered by d-glyceraldehyde 3-phosphate (GAP). Herein, we applied hydrogen-deuterium (H/D) exchange MS (HDX-MS) of full-length DXPS to provide a snapshot of the conformational dynamics of this enzyme, leading to the following conclusions. () The high sequence coverage of DXPS allowed us to monitor structural changes throughout the entire enzyme, including two segments (spanning residues 183-238 and 292-317) not observed by X-ray crystallography. () Three regions of DXPS (spanning residues 42-58, 183-199, and 278-298) near the active center displayed both EX1 (monomolecular) and EX2 (bimolecuar) H/D exchange (HDX) kinetic behavior in both ligand-free and ligand-bound states. All other peptides behaved according to the common EX2 kinetic mechanism. () The observation of conformational changes on DXPS provides support for the role of conformational dynamics in the DXPS mechanism: The closed conformation of DXPS is critical for stabilization of LThDP, whereas addition of GAP converts DXPS to the open conformation that coincides with decarboxylation of LThDP and DXP release.
Topics: Glyceraldehyde 3-Phosphate; Mass Spectrometry; Models, Molecular; Pentosephosphates; Phosphonoacetic Acid; Protein Binding; Protein Conformation; Transferases
PubMed: 28808005
DOI: 10.1073/pnas.1619981114 -
Infection Oct 2017Recurrent anogenital herpes simplex virus infections are common in patients with human immunodeficiency virus (HIV), of whom approximately 5% develop resistance to...
Recurrent anogenital herpes simplex virus infections are common in patients with human immunodeficiency virus (HIV), of whom approximately 5% develop resistance to acyclovir. We present a case of a 49-year-old man with HIV who had an 8-year history of recurrent left inguinal herpes simplex virus type 2 ulcerations. He initially responded to oral acyclovir, but developed resistance to acyclovir and eventually foscarnet. The lesion progressed to a large hypertrophic mass that required surgical excision, which led to resolution without recurrences. Our case highlights the importance of surgical excision as a treatment option in refractory herpes simplex virus anogenital infections.
Topics: Acyclovir; Antiviral Agents; Drug Resistance, Viral; Foscarnet; HIV Infections; Herpes Genitalis; Herpesvirus 2, Human; Humans; Male; Middle Aged
PubMed: 28508238
DOI: 10.1007/s15010-017-1027-y -
Journal of Virology Jul 2017Human herpesviruses 6A/B (HHV-6A/B) can integrate their viral genomes in the telomeres of human chromosomes. The viral and cellular factors contributing to HHV-6A/B...
Human herpesviruses 6A/B (HHV-6A/B) can integrate their viral genomes in the telomeres of human chromosomes. The viral and cellular factors contributing to HHV-6A/B integration remain largely unknown, mostly due to the lack of efficient and reproducible cell culture models to study HHV-6A/B integration. In this study, we characterized the HHV-6A/B integration efficiencies in several human cell lines using two different approaches. First, after a short-term infection (5 h), cells were processed for single-cell cloning and analyzed for chromosomally integrated HHV-6A/B (ciHHV-6A/B). Second, cells were infected with HHV-6A/B and allowed to grow in bulk for 4 weeks or longer and then analyzed for the presence of ciHHV-6. Using quantitative PCR (qPCR), droplet digital PCR, and fluorescent hybridization, we could demonstrate that HHV-6A/B integrated in most human cell lines tested, including telomerase-positive (HeLa, MCF-7, HCT-116, and HEK293T) and telomerase-negative cell lines (U2OS and GM847). Our results also indicate that inhibition of DNA replication, using phosphonoacetic acid, did not affect HHV-6A/B integration. Certain clones harboring ciHHV-6A/B spontaneously express viral genes and proteins. Treatment of cells with phorbol ester or histone deacetylase inhibitors triggered the expression of many viral genes, including , , and , without the production of infectious virus, suggesting that the tested stimuli were not sufficient to trigger full reactivation. In summary, both integration models yielded comparable results and should enable the identification of viral and cellular factors contributing to HHV-6A/B integration and the screening of drugs influencing viral gene expression, as well as the release of infectious HHV-6A/B from the integrated state. The analysis and understanding of HHV-6A/B genome integration into host DNA is currently limited due to the lack of reproducible and efficient viral integration systems. In the present study, we describe two quantitative cell culture viral integration systems. These systems can be used to define cellular and viral factors that play a role in HHV-6A/B integration. Furthermore, these systems will allow us to decipher the conditions resulting in virus gene expression and excision of the integrated viral genome resulting in reactivation.
Topics: Cell Culture Techniques; Cell Line; Herpesvirus 6, Human; Humans; In Situ Hybridization, Fluorescence; Real-Time Polymerase Chain Reaction; Virus Cultivation; Virus Integration
PubMed: 28468878
DOI: 10.1128/JVI.00437-17 -
Frontiers in Cellular and Infection... 2017Many microorganisms produce phosphonates, molecules characterized by stable carbon-phosphorus bonds that store phosphorus or act as antimicrobials. The role of...
Many microorganisms produce phosphonates, molecules characterized by stable carbon-phosphorus bonds that store phosphorus or act as antimicrobials. The role of phosphonates in the marine biosphere is well characterized but the role of these molecules in the intestine is poorly understood. uses its virulence factors to influence the host immune response to compete with the host and normal microflora for nutrients. cannot produce phosphonates but encodes the enzymes to use them suggesting that it is exposed to phosphonates during its life cycle. The role of phosphonates during enteric salmonellosis is unexplored. We have previously shown that , encoding a putative regulator of phosphonate metabolism, is needed for colonization in calves. Here, we report that the necessity of in colonization of the murine intestine results from multiple factors. is needed for full activation of the type-3 secretion system-1 and for optimal invasion of epithelial cells. The Δ mutant grows poorly in phosphonoacetic acid (PA) as the sole phosphorus source, but can use 2-aminoethylphosphonate. PhnA, an enzyme required for PA breakdown, is not controlled by STM3602 suggesting an additional mechanism for utilization of PA in . Typhimurium. Finally, the requirement of for intestinal colonization differs depending on the composition of the microflora. Our data suggest that has multiple regulatory targets that are necessary for survival within the microbial community in the intestine. Determination of the members of the regulon may illuminate new pathways needed for colonization of the host.
Topics: Animals; Gene Expression Regulation, Bacterial; Intestines; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Phosphonoacetic Acid; Salmonella Infections, Animal; Salmonella enterica
PubMed: 28361036
DOI: 10.3389/fcimb.2017.00069 -
Medicine Feb 2017The case reported the rapid remission of disease recurrence achieved adding foscarnet, a DNA polymerase inhibitor that interacts with fibroblast growth factor 2, to low...
RATIONALE
The case reported the rapid remission of disease recurrence achieved adding foscarnet, a DNA polymerase inhibitor that interacts with fibroblast growth factor 2, to low molecular weight heparin and sunitinib for the first time in a patient with an anaplastic thyroid cancer (ATC).
PATIENT CONCERNS
A 65-year-old woman with a multinodular goiter referred for a rapid enlargement of a nodule. Histological examination revealed an ATC with a little area of papillary thyroid cancer (PTC). The patient was resistant to selective single-target treatment.
DIAGNOSES
Immunophenotyping and gene analyses found a significant increase in FGF2 and FGFR1 expression in the primary ATC area (FGF2 = 38.2 ± 6.2% in ATC vs 34.6 ± 6.0% in the differentiated area of PTC, P < 0.05; FGFR1: 41.7 ± 6.0% in ATC vs 34.4 ± 4.2% in PTC, P < 0.001) and in metastatic neck lymph nodes (P < 0.001 vs normal control tissues). Unlike conventional imaging, F-FDG PET/CT with PERCIST 1.0 criteria promptly and quantitatively detected disease recurrence and remission before and after multitarget therapy, combining anatomic, metabolic, and functional data.
INTERVENTIONS
Foscarnet was administered given the positivity for FGF2, FGFR1 and FGFR4 in ATC. Low molecular wight heparin and Sunitinib were coadministere to limiti metastatic progression and on neck tumor masse, respectively.
OUTCOMES
The rationale for the clinical response to this innovative multitarget association with foscarnet is based on the histological and genetic finding that fibroblast growth factors and their receptor super-family are up-regulated in the primary anaplastic thyroid tumor and in the metastatic lymph node of our patient.
LESSONS
We propose that fibroblast growth factors and their receptor super-family play a key role as potential therapeutic targets in anaplastic thyroid cancer and the positive relevance of this suggestion for patient care, especially for an individualized management.
Topics: Aged; Antineoplastic Combined Chemotherapy Protocols; Antiviral Agents; Female; Fluorodeoxyglucose F18; Foscarnet; Humans; Positron Emission Tomography Computed Tomography; Thyroid Carcinoma, Anaplastic; Thyroid Neoplasms
PubMed: 28178124
DOI: 10.1097/MD.0000000000005621 -
Virus Research Apr 2017Vaccinia virus is the prototypic poxvirus. The 192 kilobase double-stranded DNA viral genome encodes most if not all of the viral replication machinery. The vaccinia... (Review)
Review
Vaccinia virus is the prototypic poxvirus. The 192 kilobase double-stranded DNA viral genome encodes most if not all of the viral replication machinery. The vaccinia virus DNA polymerase is encoded by the E9L gene. Sequence analysis indicates that E9 is a member of the B family of replicative polymerases. The enzyme has both polymerase and 3'-5' exonuclease activities, both of which are essential to support viral replication. Genetic analysis of E9 has identified residues and motifs whose alteration can confer temperature-sensitivity, drug resistance (phosphonoacetic acid, aphidicolin, cytosine arabinsode, cidofovir) or altered fidelity. The polymerase is involved both in DNA replication and in recombination. Although inherently distributive, E9 gains processivity by interacting in a 1:1 stoichiometry with a heterodimer of the A20 and D4 proteins. A20 binds to both E9 and D4 and serves as a bridge within the holoenzyme. The A20/D4 heterodimer has been purified and can confer processivity on purified E9. The interaction of A20 with D4 is mediated by the N'-terminus of A20. The D4 protein is an enzymatically active uracil DNA glycosylase. The DNA-scanning activity of D4 is proposed to keep the holoenzyme tethered to the DNA template but allow polymerase translocation. The crystal structure of D4, alone and in complex with A20 and/or DNA has been solved. Screens for low molecular weight compounds that interrupt the A20/D4 interface have yielded hits that disrupt processive DNA synthesis in vitro and/or inhibit plaque formation. The observation that an active DNA repair enzyme is an integral part of the holoenzyme suggests that DNA replication and repair may be coupled.
Topics: DNA, Viral; DNA-Directed DNA Polymerase; Protein Binding; Protein Multimerization; Recombination, Genetic; Uracil-DNA Glycosidase; Vaccinia virus; Viral Proteins; Virus Replication
PubMed: 28159613
DOI: 10.1016/j.virusres.2017.01.027 -
Internal Medicine (Tokyo, Japan) 2017Infections of the central nervous system (CNS) with varicella zoster virus (VZV) is a rare occurrence after allogeneic hematopoietic stem cell transplantation. We herein...
Infections of the central nervous system (CNS) with varicella zoster virus (VZV) is a rare occurrence after allogeneic hematopoietic stem cell transplantation. We herein report a case of VZV meningitis, radiculitis and myelitis that developed 8 months after cord blood transplantation, shortly after the cessation of cyclosporine and low-dose acyclovir. Although treatment with acyclovir did not achieve a satisfactory response, the patient was successfully treated with foscarnet. Our report indicates that VZV infection should be considered in allo-hematopoietic stem cell transplantation (HSCT) patients with CNS symptoms and that foscarnet may be effective for the treatment of acyclovir-resistant VZV infections of the CNS. The development of optimal prophylactic strategies and vaccination schedules may eradicate post-transplant VZV disease.
Topics: Acyclovir; Antiviral Agents; Foscarnet; Graft vs Host Disease; Hematopoietic Stem Cell Transplantation; Herpes Zoster; Herpesvirus 3, Human; Humans; Meningitis, Viral; Transplantation, Homologous
PubMed: 28154282
DOI: 10.2169/internalmedicine.56.6930 -
ACS Synthetic Biology Feb 2017The activation of silent natural product gene clusters is a synthetic biology problem of great interest. As the rate at which gene clusters are identified outpaces the...
The activation of silent natural product gene clusters is a synthetic biology problem of great interest. As the rate at which gene clusters are identified outpaces the discovery rate of new molecules, this unknown chemical space is rapidly growing, as too are the rewards for developing technologies to exploit it. One class of natural products that has been underrepresented is phosphonic acids, which have important medical and agricultural uses. Hundreds of phosphonic acid biosynthetic gene clusters have been identified encoding for unknown molecules. Although methods exist to elicit secondary metabolite gene clusters in native hosts, they require the strain to be amenable to genetic manipulation. One method to circumvent this is pathway refactoring, which we implemented in an effort to discover new phosphonic acids from a gene cluster from Streptomyces sp. strain NRRL F-525. By reengineering this cluster for expression in the production host Streptomyces lividans, utility of refactoring is demonstrated with the isolation of a novel phosphonic acid, O-phosphonoacetic acid serine, and the characterization of its biosynthesis. In addition, a new biosynthetic branch point is identified with a phosphonoacetaldehyde dehydrogenase, which was used to identify additional phosphonic acid gene clusters that share phosphonoacetic acid as an intermediate.
Topics: Biological Products; Hydrolases; Multigene Family; Phosphonoacetic Acid; Phosphorous Acids; Streptomyces; Synthetic Biology
PubMed: 28103011
DOI: 10.1021/acssynbio.6b00299 -
Clinical Microbiology and Infection :... May 2017Staphylococcus lugdunensis is a coagulase-negative staphylococcus that displays an unusually high virulence rate close to that of Staphylococcus aureus. It also shares...
OBJECTIVE
Staphylococcus lugdunensis is a coagulase-negative staphylococcus that displays an unusually high virulence rate close to that of Staphylococcus aureus. It also shares phenotypic properties with S. aureus and several studies found putative virulence factors. The objective of the study was to describe the clinical manifestations of S. lugdunensis infections and investigate putative virulence factors.
METHOD
We conducted a prospective study from November 2013 to March 2016 at the University Hospital of Strasbourg. Putative virulence factors were investigated by clumping factor detection, screening for proteolytic activity, and sequence analysis using tandem nano-liquid chromatography-mass spectrometry.
RESULTS
In total, 347 positive samples for S. lugdunensis were collected, of which 129 (37.2%) were from confirmed cases of S. lugdunensis infection. Eighty-one of these 129 patients were included in the study. Bone and prosthetic joints (PJI) were the most frequent sites of infection (n=28; 34.6%) followed by skin and soft tissues (n=23; 28.4%). We identified and purified a novel protease secreted by 50 samples (61.7%), most frequently associated with samples from deep infections and PJI (pr 0.97 and pr 0.91, respectively). Protease peptide sequencing by nano-liquid chromatography-mass spectrometry revealed a novel protease bearing 62.42% identity with ShpI, a metalloprotease secreted by Staphylococcus hyicus.
CONCLUSION
This study confirms the pathogenicity of S. lugdunensis, particularly in bone and PJI. We also identified a novel metalloprotease called lugdulysin that may contribute to virulence.
Topics: Adult; Aged; Aged, 80 and over; Amino Acid Sequence; Aminoglycosides; Base Sequence; DNA, Bacterial; Drug Resistance, Multiple, Bacterial; Erythromycin; Female; Fluoroquinolones; Follow-Up Studies; Fosfomycin; Fusidic Acid; Humans; Male; Metalloproteases; Methicillin; Middle Aged; Phosphonoacetic Acid; Prospective Studies; Sequence Analysis, DNA; Staphylococcal Infections; Staphylococcus lugdunensis; Vancomycin; Virulence Factors
PubMed: 28017792
DOI: 10.1016/j.cmi.2016.12.018