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Microbiology
Rapid single-cell detection and identification of pathogens by using surface-enhanced Raman spectroscopy
Apr 21, 2017   The Analyst
Dina NE, Zhou H, Colniţă A, Leopold N, Szoke-Nagy T, Coman C, Haisch C
Rapid single-cell detection and identification of pathogens by using surface-enhanced Raman spectroscopy
Apr 21, 2017
The Analyst
For the successful treatment of infections, real-time analysis and enhanced multiplex capacity, sensitivity and cost-effectiveness of the developed detection method are critical. In this work, surface-enhanced Raman scattering (SERS) was employed with the final aim of identification and discrimination of pathogenic bacteria, based on their detected SERS fingerprint at the single-cell level. Several genera of bacteria that are found in most of the isolated infections in bacteraemia were successfully identified in less than 5 minutes without the use of antibodies or other specific receptors. The key element of the SERS direct detection platform is the SERS substrate, which combines easy production at low costs with a high enhancement enabling single-cell detection. The innovative approach of detection required the in situ synthesis of silver nanoparticles (NPs), ensuring an intimate contact with the bacterial membrane. This protocol provided a good reproducibility of the single-cell SERS spectra and was successfully applied both on Gram-negative and Gram-positive microorganisms (E. coli, M. morganii, E. lactis, L. casei). Thus, a label-free SERS-based biosensor for pathogen detection was developed with low costs, minimal sample preparation, high-accuracy and a very short analysis time of less than 5 min, which is crucial for infection diagnosis.
Reduction of pulmonary toxicity of metal oxide nanoparticles by phosphonate-based surface passivation
Apr 22, 2017   Particle And Fibre Toxicology
Cai X, Lee A, Ji Z, Huang C, Chang CH, Wang X, Liao YP, Xia T, Li R
Reduction of pulmonary toxicity of metal oxide nanoparticles by phosphonate-based surface passivation
Apr 22, 2017
Particle And Fibre Toxicology
The wide application of engineered nanoparticles has induced increasing exposure to humans and environment, which led to substantial concerns on their biosafety. Some metal oxides (MOx) have shown severe toxicity in cells and animals, thus safe designs of MOx with reduced hazard potential are desired. Currently, there is a lack of a simple yet effective safe design approach for the toxic MOx. In this study, we determined the key physicochemical properties of MOx that lead to cytotoxicity and explored a safe design approach for toxic MOx by modifying their hazard properties. THP-1 and BEAS-2B cells were exposed to 0-200 μg/mL MOx for 24 h, we found some toxic MOx including CoO, CuO, Ni2O3 and Co3O4, could induce reactive oxygen species (ROS) generation and cell death due to the toxic ion shedding and/or oxidative stress generation from the active surface of MOx internalized into lysosomes. We thus hypothesized that surface passivation could reduce or eliminate the toxicity of MOx. We experimented with a series of surface coating molecules and discovered that ethylenediamine tetra (methylene phosphonic acid) (EDTMP) could form stable hexadentate coordination with MOx. The coating layer can effectively reduce the surface activity of MOx with 85-99% decrease of oxidative potential, and 65-98% decrease of ion shedding. The EDTMP coated MOx show negligible ROS generation and cell death in THP-1 and BEAS-2B cells. The protective effect of EDTMP coating was further validated in mouse lungs exposed to 2 mg/kg MOx by oropharyngeal aspiration. After 40 h exposure, EDTMP coated MOx show significant decreases of neutrophil counts, lactate dehydrogenase (LDH) release, MCP-1, LIX and IL-6 in bronchoalveolar lavage fluid (BALF), compared to uncoated particles. The haematoxylin and eosin (H&E) staining results of lung tissue also show EDTMP coating could significantly reduce the pulmonary inflammation of MOx. The surface reactivity of MOx including ion shedding and oxidative potential is the dominated physicochemical property that is responsible for the cytotoxicity induced by MOx. EDTMP coating could passivate the surface of MOx, reduce their cytotoxicity and pulmonary hazard effects. This coating would be an effective safe design approach for a broad spectrum of toxic MOx, which will facilitate the safe use of MOx in commercial nanoproducts.
A humanized mouse-based HIV-1 viral outgrowth assay with higher sensitivity than in vitro qVOA in detecting latently infected cells from individuals on ART with undetectable viral loads
Apr 22, 2017   Virology
Charlins P, Schmitt K, Remling-Mulder L, Hogan LE, Hanhauser E, Hobbs KS, Hecht F, Deeks SG, Henrich TJ, Akkina R
A humanized mouse-based HIV-1 viral outgrowth assay with higher sensitivity than in vitro qVOA in detecting latently infected cells from individuals on ART with undetectable viral loads
Apr 22, 2017
Virology
Assays that can verify full viral eradication are essential in the context of achieving a cure for HIV/AIDS. In vitro quantitative viral out growth assays (qVOA) are currently the gold standard for measuring latent HIV-1 but these assays often fail to detect very low levels of replication-competent virus. Here we investigated an alternative in vivo approach for sensitive viral detection using humanized mice (hmVOA). Peripheral blood CD4+ T cell samples from HIV subjects on stable ART with undetectable viral loads by RT-PCR were first assayed by in vitro qVOA. Corresponding patient samples in which no virus was detected by qVOA were injected into humanized mice to allow viral outgrowth. Of the five qVOA virus negative samples, four gave positive viral outgrowth in the hmVOA assay suggesting that it is more sensitive in detecting latent HIV-1. Copyright © 2017. Published by Elsevier Inc.
Structure of the S1 subunit C-terminal domain from bat-derived coronavirus HKU5 spike protein
Apr 22, 2017   Virology
Han X, Qi J, Song H, Wang Q, Zhang Y, Wu Y, Lu G, Yuen KY, Shi Y, Gao GF
Structure of the S1 subunit C-terminal domain from bat-derived coronavirus HKU5 spike protein
Apr 22, 2017
Virology
Accumulating evidence indicates that MERS-CoV originated from bat coronaviruses (BatCoVs). Previously, we demonstrated that both MERS-CoV and BatCoV HKU4 use CD26 as a receptor, but how the BatCoVs evolved to bind CD26 is an intriguing question. Here, we solved the crystal structure of the S1 subunit C-terminal domain of HKU5 (HKU5-CTD), another BatCoV that is phylogenetically related to MERS-CoV but cannot bind to CD26. We observed that the conserved core subdomain and those of other betacoronaviruses (betaCoVs) have a similar topology of the external subdomain, indicating the same ancestor of lineage C betaCoVs. However, two deletions in two respective loops located in HKU5-CTD result in conformational variations in CD26-binding interface and are responsible for the non-binding of HKU5-CTD to CD26. Combined with sequence variation in the HKU5-CTD receptor binding interface, we propose the necessity for surveilling the mutation in BatCoV HKU5 spike protein in case of bat-to-human interspecies transmission. Copyright © 2017 Elsevier Inc. All rights reserved.
Structure-function analysis of the DNA-binding domain of a transmembrane transcriptional activator
Apr 22, 2017   Scientific Reports
Schlundt A, Buchner S, Janowski R, Heydenreich T, Heermann R, Lassak J, Geerlof A, Stehle R, Niessing D, Jung K, Sattler M
Structure-function analysis of the DNA-binding domain of a transmembrane transcriptional activator
Apr 22, 2017
Scientific Reports
The transmembrane DNA-binding protein CadC of E. coli, a representative of the ToxR-like receptor family, combines input and effector domains for signal sensing and transcriptional activation, respectively, in a single protein, thus representing one of the simplest signalling systems. At acidic pH in a lysine-rich environment, CadC activates the transcription of the cadBA operon through recruitment of the RNA polymerase (RNAP) to the two cadBA promoter sites, Cad1 and Cad2, which are directly bound by CadC. However, the molecular details for its interaction with DNA have remained elusive. Here, we present the crystal structure of the CadC DNA-binding domain (DBD) and show that it adopts a winged helix-turn-helix fold. The interaction with the cadBA promoter site Cad1 is studied by using nuclear magnetic resonance (NMR) spectroscopy, biophysical methods and functional assays and reveals a preference for AT-rich regions. By mutational analysis we identify amino acids within the CadC DBD that are crucial for DNA-binding and functional activity. Experimentally derived structural models of the CadC-DNA complex indicate that the CadC DBD employs mainly non-sequence-specific over a few specific contacts. Our data provide molecular insights into the CadC-DNA interaction and suggest how CadC dimerization may provide high-affinity binding to the Cad1 promoter.
Do Memory CD4 T Cells Keep Their Cell-Type Programming: Plasticity versus Fate Commitment? T-Cell Heterogeneity, Plasticity, and Selection in Humans
Apr 22, 2017   Cold Spring Harbor Perspectives In Biology
Sallusto F, Cassotta A, Hoces D, Foglierini M, Lanzavecchia A
Do Memory CD4 T Cells Keep Their Cell-Type Programming: Plasticity versus Fate Commitment? T-Cell Heterogeneity, Plasticity, and Selection in Humans
Apr 22, 2017
Cold Spring Harbor Perspectives In Biology
The wide range of effector and memory T cells is instrumental for immune regulation and tailored mechanisms of protection against pathogens. Here, we will focus on human CD4 T cells and discuss T-cell plasticity and intraclonal diversification in the context of a progressive and selective model of CD4 T-cell differentiation. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.
Do the Microbiota Influence Vaccines and Protective Immunity to Pathogens? Engaging Our Endogenous Adjuvants
Apr 22, 2017   Cold Spring Harbor Perspectives In Biology
Collins N, Belkaid Y
Do the Microbiota Influence Vaccines and Protective Immunity to Pathogens? Engaging Our Endogenous Adjuvants
Apr 22, 2017
Cold Spring Harbor Perspectives In Biology
The reliance of the immune system on constitutive microbial stimulation support the idea that both responsiveness to vaccines and vaccine design need to be considered in the context of host-microbiota interactions. Manipulation of microbe function or composition via diet alteration or microbiota engraftment may soon become a viable approach to control immunity and, as such, vaccine responses. Learning from our endogenous original adjuvants could be critical in overcoming the enormous hurdle of vaccine design against the numerous pathogens that cause chronic infection. Going forward, rationally designed vaccines that take advantage of the inherent adjuvant properties of the microbiota could have a major impact on the prevention of disease. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.
Acute Hepatopancreatic Necrosis Disease (AHPND)-causing Vibrio parahaemolyticus strains maintain an antibacterial Type VI Secretion System with versatile effector repertoires
Apr 22, 2017   Applied And Environmental Microbiology
Li P, Kinch LN, Ray A, Dalia AB, Cong Q, Nunan LM, Camilli A, Grishin NV, Salomon D, Orth K
Acute Hepatopancreatic Necrosis Disease (AHPND)-causing Vibrio parahaemolyticus strains maintain an antibacterial Type VI Secretion System with versatile effector repertoires
Apr 22, 2017
Applied And Environmental Microbiology
Acute hepatopancreatic necrosis disease (AHPND) is a newly emerging shrimp disease that has severely damaged the global shrimp industry. AHPND is caused by toxic strains of Vibrio parahaemolyticus (V. parahaemolyticus) that have acquired a "selfish plasmid" encoding the deadly binary toxins PirAvp/PirBvp To better understand the repertoire of virulence factors in AHPND-causing V. parahaemolyticus, we conducted a comparative analysis using genome sequences of the clinical strain RIMD2210633, environmental non-AHPND and toxic AHPND isolates of V. parahaemolyticus Interestingly, we found that all of the AHPND strains, but none of the non-AHPND strains, harbor the antibacterial type VI secretion system 1 (T6SS1) which we previously identified and characterized in the clinical isolate RIMD2210633. This finding suggests that the acquisition of this T6SS might confer AHPND-causing V. parahaemolyticus a fitness advantage over competing bacteria and facilitate shrimp infection. Additionally, we find highly dynamic effector loci in the T6SS1 of AHPND-causing strains, leading to diverse effector repertoires. Our discovery provides novel insights into AHPND-causing pathogens and reveals a potential target for disease control.Importance Acute hepatopancreatic necrosis disease (AHPND) is a serious disease that has caused severe damage and significant financial losses to the global shrimp industry. To better understand and prevent this shrimp disease, it is essential to thoroughly characterize its causative agent, Vibrio paraehaemolyticus (V. parahaemolyticus). Although the plasmid-encoded binary toxins PirAvp/PirBvp have been shown to be the primary cause of AHPND, it remains unknown whether other virulent factors are commonly present in V. parahaemolyticus and might play important roles during shrimp infection. Here, we analyzed the genome sequences of clinical, non-AHPND and AHPND strains to characterize their repertoires of key virulence determinants. Our studies reveal that an antibacterial type VI secretion system is associated with the AHPND strains and differentiates them from non-AHPND strains, similar to the PirA/PirB toxins. We propose that T6SS1 provides a selective advantage during shrimp infections. Copyright © 2017 American Society for Microbiology.
In vitro fermentation behaviors of fucosylated chondroitin sulfate from Pearsonothuria graeffei by human gut microflora
Apr 22, 2017   International Journal Of Biological Macromolecules
Wei CY, Liao NB, Zhang Y, Ye XQ, Li S, Hu YQ, Liu DH, Linhardt RJ, Wang X, Chen SG
In vitro fermentation behaviors of fucosylated chondroitin sulfate from Pearsonothuria graeffei by human gut microflora
Apr 22, 2017
International Journal Of Biological Macromolecules
A fucosylated chondroitin sulfate (FCS-pg) with highly repeated structure from Pearsonothuria graeffei was subjected to a in vitro fermentation model to investigate its fermentability and effects on human gut microflora. High performance liquid chromatography (HPLC) measurement found FCS-pg can be fermented to short chain fatty acids (SCFAs) by gut microflora from partial human fecal samples. 16S rRNA gene-based polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE) profiling and real-time quantitative PCR analysis showed that FCS-pg mainly increased the proportions of Clostridium cluster XI, Bacteriodes prevotella group, Bifidobacterium genus, Clostridium cluster I and Clostridium cluster XIVab, whereas the numbers of the Enterobacteriaceae and Lactobacillus decreased. These results indicated that FCS-pg is mainly fermented by Bacteroides, Bifidobacterium and Clostridium. It increased the content of probiotics bacteria in achieving health-enhancing effect, was slightly different than most sulfated polysaccharides from marine animals. The current study provides useful new information on the mechanism of absorption and functional activity on FCS-pg within the gastrointestinal tract of the human body. Copyright © 2017. Published by Elsevier B.V.
Molecular cloning and functional characterization of duck nucleotide-binding oligomerization domain 1 (NOD1)
Apr 22, 2017   Developmental And Comparative Immunology
Li H, Jin H, Li Y, Liu D, Foda MF, Jiang Y, Luo R
Molecular cloning and functional characterization of duck nucleotide-binding oligomerization domain 1 (NOD1)
Apr 22, 2017
Developmental And Comparative Immunology
Nucleotide-binding oligomerization domain 1 (NOD1) is an imperative cytoplasmic pattern recognition receptor (PRR) and considered as a key member of the NOD-like receptor (NLR) family which plays a critical role in innate immunity through sensing microbial components derived from bacterial peptidoglycan. In the current study, the full-length of duck NOD1 (duNOD1) cDNA from duck embryo fibroblasts (DEFs) was cloned. Multiple sequence alignment and phylogenetic analysis demonstrated that duNOD1 exhibited a strong evolutionary relationship with chicken and rock pigeon NOD1. Tissue-specific expression analysis showed that duNOD1 was widely distributed in various organs, with the highest expression observed in the liver. Furthermore, duNOD1 overexpression induced NF-κB activation in DEFs and the CARD domain is crucial for duNOD1-mediated NF-κB activation. In addition, silencing the duNOD1 decreased the activity of NF-κB in DEFs stimulated by iE-DAP. Overexpression of duNOD1 significantly increased the expression of TNF-α, IL-6, and RANTES in DEFs. These findings highlight the crucial role of duNOD1 as an intracellular sensor in duck innate immune system. Copyright © 2017. Published by Elsevier Ltd.
How well does transfer of bacterial pathogens by culture swabs correlate with transfer by hands?
Apr 22, 2017   American Journal Of Infection Control
Kanwar A, Mana TSC, Cadnum JL, Alhmidi H, Koganti S, Donskey CJ
How well does transfer of bacterial pathogens by culture swabs correlate with transfer by hands?
Apr 22, 2017
American Journal Of Infection Control
In laboratory testing and in isolation rooms, pickup and transfer of health care-associated pathogens by premoistened rayon swabs correlated well with pickup and transfer by bare hands or moistened gloves. These results suggest that swab cultures provide a useful surrogate indicator of the risk for pathogen pickup and transfer by hands. Published by Elsevier Inc.
Enhanced neuroinvasion by smaller, soluble prions
Apr 22, 2017   Acta Neuropathologica Communications
Bett C, Lawrence J, Kurt TD, Orru C, Aguilar-Calvo P, Kincaid AE, Surewicz WK, Caughey B, Wu C, Sigurdson CJ
Enhanced neuroinvasion by smaller, soluble prions
Apr 22, 2017
Acta Neuropathologica Communications
Infectious prion aggregates can propagate from extraneural sites into the brain with remarkable efficiency, likely transported via peripheral nerves. Yet not all prions spread into the brain, and the physical properties of a prion that is capable of transit within neurons remain unclear. We hypothesized that small, diffusible aggregates spread into the CNS via peripheral nerves. Here we used a structurally diverse panel of prion strains to analyze how the prion conformation impacts transit into the brain. Two prion strains form fibrils visible ultrastructurally in the brain in situ, whereas three strains form diffuse, subfibrillar prion deposits and no visible fibrils. The subfibrillar strains had significantly higher levels of soluble prion aggregates than the fibrillar strains. Primary neurons internalized both the subfibrillar and fibril-forming prion strains by macropinocytosis, and both strain types were transported from the axon terminal to the cell body in vitro. However in mice, only the predominantly soluble, subfibrillar prions, and not the fibrillar prions, were efficiently transported from the tongue to the brain. Sonicating a fibrillar prion strain increased the solubility and enabled prions to spread into the brain in mice, as evident by a 40% increase in the attack rate, indicating that an increase in smaller particles enhances prion neuroinvasion. Our data suggest that the small, highly soluble prion particles have a higher capacity for transport via nerves. These findings help explain how prions that predominantly assemble into subfibrillar states can more effectively traverse into and out of the CNS, and suggest that promoting fibril assembly may slow the neuron-to-neuron spread of protein aggregates.
Tunable Expression Tools Enable Single-Cell Strain Distinction in the Gut Microbiome
Apr 21, 2017   Cell
Whitaker WR, Shepherd ES, Sonnenburg JL
Tunable Expression Tools Enable Single-Cell Strain Distinction in the Gut Microbiome
Apr 21, 2017
Cell
Applying synthetic biology to engineer gut-resident microbes provides new avenues to investigate microbe-host interactions, perform diagnostics, and deliver therapeutics. Here, we describe a platform for engineering Bacteroides, the most abundant genus in the Western microbiota, which includes a process for high-throughput strain modification. We have identified a novel phage promoter and translational tuning strategy and achieved an unprecedented level of expression that enables imaging of fluorescent-protein-expressing Bacteroides stably colonizing the mouse gut. A detailed characterization of the phage promoter has provided a set of constitutive promoters that span over four logs of strength without detectable fitness burden within the gut over 14 days. These promoters function predictably over a 1,000,000-fold expression range in phylogenetically diverse Bacteroides species. With these promoters, unique fluorescent signatures were encoded to allow differentiation of six species within the gut. Fluorescent protein-based differentiation of isogenic strains revealed that priority of gut colonization determines colonic crypt occupancy. Copyright © 2017 Elsevier Inc. All rights reserved.
A no film slot blot for the detection of developing P. falciparum oocysts in mosquitoes
Apr 21, 2017   PloS One
Grabias B, Verma N, Zheng H, Tripathi AK, Mlambo G, Morin MJ, Locke E, Kumar S
A no film slot blot for the detection of developing P. falciparum oocysts in mosquitoes
Apr 21, 2017
PloS One
Non-microscopy-based assays for sensitive and rapid detection of Plasmodium infection in mosquitoes are needed to allow rapid and high throughput measurement of transmission intensity and malaria control program effectiveness. Here, we report on a modified enhanced chemiluminescence-based slot blot assay for detection of Plasmodium falciparum (Pf) circumsporozite protein (PfCSP) expressed on parasite oocysts developing inside the mosquito midgut. This modified assay has several novel features that include eliminating the need for exposure to autoradiography (AR) film, as well as utilizing a novel high affinity anti-CSP antibody, and optimizing assay procedures resulting in significant reduction in the time required to perform the assay. The chemiluminescent signal for the detection of PfCSP in mosquito samples was captured digitally utilizing the C-Digit blot scanner that, allowed the detection of 0.01 pg of recombinant P. falciparum CSP and as few as 0.02 P. falciparum oocysts in a little over two hours. The earlier ECL-SB detected rCSP and oocysts and took approximately 5 h to perform. Whole mosquito lysates from both high and low prevalence-infected mosquito populations were prepared and evaluated for PfCSP detection on the ECL-SB by both AR film and digital data capture and analysis. There was a 100% agreement between the AR film and the C-Digit scanner methods for PfCSP detection in randomly sampled mosquitoes. This novel "No Film" Slot Blot assay obviates the need for AR film exposure and development and significantly reduces the assay time enabling widespread use in field settings.
Nuclear Proximity of Mtr4 to RNA Exosome Restricts DNA Mutational Asymmetry
Apr 21, 2017   Cell
Lim J, Giri PK, Kazadi D, Laffleur B, Zhang W,   . . . . . .   , Rothschild G, Cogné M, Pinaud E, Deng H, Basu U
Nuclear Proximity of Mtr4 to RNA Exosome Restricts DNA Mutational Asymmetry
Apr 21, 2017
Cell
The distribution of sense and antisense strand DNA mutations on transcribed duplex DNA contributes to the development of immune and neural systems along with the progression of cancer. Because developmentally matured B cells undergo biologically programmed strand-specific DNA mutagenesis at focal DNA/RNA hybrid structures, they make a convenient system to investigate strand-specific mutagenesis mechanisms. We demonstrate that the sense and antisense strand DNA mutagenesis at the immunoglobulin heavy chain locus and some other regions of the B cell genome depends upon localized RNA processing protein complex formation in the nucleus. Both the physical proximity and coupled activities of RNA helicase Mtr4 (and senataxin) with the noncoding RNA processing function of RNA exosome determine the strand-specific distribution of DNA mutations. Our study suggests that strand-specific DNA mutagenesis-associated mechanisms will play major roles in other undiscovered aspects of organismic development. Copyright © 2017 Elsevier Inc. All rights reserved.
Spontaneous Chitin Accumulation in Airways and Age-Related Fibrotic Lung Disease
Apr 21, 2017   Cell
Van Dyken SJ, Liang HE, Naikawadi RP, Woodruff PG, Wolters PJ, Erle DJ, Locksley RM
Spontaneous Chitin Accumulation in Airways and Age-Related Fibrotic Lung Disease
Apr 21, 2017
Cell
The environmentally widespread polysaccharide chitin is degraded and recycled by ubiquitous bacterial and fungal chitinases. Although vertebrates express active chitinases from evolutionarily conserved loci, their role in mammalian physiology is unclear. We show that distinct lung epithelial cells secrete acidic mammalian chitinase (AMCase), which is required for airway chitinase activity. AMCase-deficient mice exhibit premature morbidity and mortality, concomitant with accumulation of environmentally derived chitin polymers in the airways and expression of pro-fibrotic cytokines. Over time, these mice develop spontaneous pulmonary fibrosis, which is ameliorated by restoration of lung chitinase activity by genetic or therapeutic approaches. AMCase-deficient epithelial cells express fibrosis-associated gene sets linked with cell stress pathways. Mice with lung fibrosis due to telomere dysfunction and humans with interstitial lung disease also accumulate excess chitin polymers in their airways. These data suggest that altered chitin clearance could exacerbate fibrogenic pathways in the setting of lung diseases characterized by epithelial cell dysfunction. Copyright © 2017 Elsevier Inc. All rights reserved.
Host-Microbe Co-metabolism Dictates Cancer Drug Efficacy in C. elegans
Apr 21, 2017   Cell
Scott TA, Quintaneiro LM, Norvaisas P, Lui PP, Wilson MP,   . . . . . .   , Velagapudi V, Mills PB, Typas A, Greene NDE, Cabreiro F
Host-Microbe Co-metabolism Dictates Cancer Drug Efficacy in C. elegans
Apr 21, 2017
Cell
Fluoropyrimidines are the first-line treatment for colorectal cancer, but their efficacy is highly variable between patients. We queried whether gut microbes, a known source of inter-individual variability, impacted drug efficacy. Combining two tractable genetic models, the bacterium E. coli and the nematode C. elegans, we performed three-way high-throughput screens that unraveled the complexity underlying host-microbe-drug interactions. We report that microbes can bolster or suppress the effects of fluoropyrimidines through metabolic drug interconversion involving bacterial vitamin B6, B9, and ribonucleotide metabolism. Also, disturbances in bacterial deoxynucleotide pools amplify 5-FU-induced autophagy and cell death in host cells, an effect regulated by the nucleoside diphosphate kinase ndk-1. Our data suggest a two-way bacterial mediation of fluoropyrimidine effects on host metabolism, which contributes to drug efficacy. These findings highlight the potential therapeutic power of manipulating intestinal microbiota to ensure host metabolic health and treat disease. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.
Reducing Recurrence of C. difficile Infection
Apr 21, 2017   Cell
Dieterle MG, Young VB
Reducing Recurrence of C. difficile Infection
Apr 21, 2017
Cell
Clostridium difficile infection (CDI) is facilitated by alteration of the microbiome following antibiotic administration. Antimicrobial therapy directed against the pathogen can treat CDI. Unfortunately, ∼20% of successfully treated patients will suffer recurrence. Bezlotoxumab, a human monoclonal antibody, binds to C. difficile toxin B (TcdB), reducing recurrence presumably by limiting epithelial damage and facilitating microbiome recovery. Copyright © 2017 Elsevier Inc. All rights reserved.
Structural Basis for Guide RNA Processing and Seed-Dependent DNA Targeting by CRISPR-Cas12a
Apr 21, 2017   Molecular Cell
Swarts DC, van der Oost J, Jinek M
Structural Basis for Guide RNA Processing and Seed-Dependent DNA Targeting by CRISPR-Cas12a
Apr 21, 2017
Molecular Cell
The CRISPR-associated protein Cas12a (Cpf1), which has been repurposed for genome editing, possesses two distinct nuclease activities: endoribonuclease activity for processing its own guide RNAs and RNA-guided DNase activity for target DNA cleavage. To elucidate the molecular basis of both activities, we determined crystal structures of Francisella novicida Cas12a bound to guide RNA and in complex with an R-loop formed by a non-cleavable guide RNA precursor and a full-length target DNA. Corroborated by biochemical experiments, these structures reveal the mechanisms of guide RNA processing and pre-ordering of the seed sequence in the guide RNA that primes Cas12a for target DNA binding. Furthermore, the R-loop complex structure reveals the strand displacement mechanism that facilitates guide-target hybridization and suggests a mechanism for double-stranded DNA cleavage involving a single active site. Together, these insights advance our mechanistic understanding of Cas12a enzymes and may contribute to further development of genome editing technologies. Copyright © 2017 Elsevier Inc. All rights reserved.
An Escherichia coli expression model reveals the species-specific function of FtsA from Neisseria gonorrhoeae in cell division
Apr 21, 2017   FEMS Microbiology Letters
Zou Y, Li Y, Ekanayake SB, Dillon JR
An Escherichia coli expression model reveals the species-specific function of FtsA from Neisseria gonorrhoeae in cell division
Apr 21, 2017
FEMS Microbiology Letters
Escherichia coli (Ec) has been used to study the function of cell division proteins from different microorganisms, especially when genetic tools are limited for studying these proteins in their native hosts. The expression of ftsA from N. gonorrhoeae (Ng) disrupted cell division in E. coli resulting in a significant increase in cell length. In some cells, FtsANg localized to the division site and the poles of E. coli cells, but the majority of cells showed no specifical localization. FtsANg did not complement an E. coli ftsA mutant strain. Bacterial two-hybrid and GST pull-down assays indicated that FtsANg interacted with FtsNEc, but no other cell division proteins from E. coli. This interaction was mediated through the 2A and 2B subdomains of FtsANg. This evidence suggests that the function of FtsANg is species-specific. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
Diabetes mellitus as the major risk factor for mucormycosis in Mexico: Epidemiology, diagnosis, and outcomes of reported cases
Apr 21, 2017   Medical Mycology
Corzo-León DE, Chora-Hernández LD, Rodríguez-Zulueta AP, Walsh TJ
Diabetes mellitus as the major risk factor for mucormycosis in Mexico: Epidemiology, diagnosis, and outcomes of reported cases
Apr 21, 2017
Medical Mycology
Mucormycosis is an emerging infectious disease with high rates of associated mortality and morbidity. Little is known about the characteristics of mucormycosis or entomophthoromycosis occurring in Mexico. A search strategy was performed of literature published in journals found in available databases and theses published online at Universidad Nacional Autónoma de México (UNAM) library website reporting clinical cases or clinical case series of mucormycosis and entomophthoromycosis occurring in Mexico between 1982 and 2016. Among the 418 cases identified, 72% were diabetic patients, and sinusitis accounted for 75% of the reported cases. Diabetes mellitus was not a risk factor for entomophthoromycosis. Mortality rate was 51% (125/244). Rhizopus species were the most frequent isolates (59%, 148/250). Amphotericin B deoxycholate was used in 89% of cases (204/227), while surgery and antifungal management as combined treatment was used in 90% (172/191). In diabetic individuals, this combined treatment approach was associated with a higher probability of survival (95% vs 66%, OR = 0.1, 95% CI, 0.02-0.43' P = .002). The most common complications were associated with nephrotoxicity and prolonged hospitalization due to IV antifungal therapy. An algorithm is proposed to establish an early diagnosis of rhino-orbital cerebral (ROC) mucormycosis based on standardized identification of warning signs and symptoms and performing an early direct microbiological exam and histopathological identification through a multidisciplinary medical and surgical team. In summary, diabetes mellitus was the most common risk factor for mucormycosis in Mexico; combined antifungal therapy and surgery in ROC mucormycosis significantly improved survival. © The Author 2017. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
Repurposing QuantiFERON for detection of neutralizing IFN-γ autoantibodies in patients with nontuberculous mycobacterial infections
Apr 21, 2017   Clinical Infectious Diseases : An Official Publication Of The Infectious Diseases Society Of America
Suárez I, Lehmann C, Gruell H, Graeb J, Kochanek M, Fätkenheuer G, Plum G, van Wengen A, van de Vosse E, Hartmann P, Hanitsch LG, Rybniker J
Repurposing QuantiFERON for detection of neutralizing IFN-γ autoantibodies in patients with nontuberculous mycobacterial infections
Apr 21, 2017
Clinical Infectious Diseases : An Official Publication Of The Infectious Diseases Society Of America
Nontuberculous mycobacterial (NTM) infections due to autoantibodies targeting IFN-γ are an emerging medical problem. However, case finding is hampered due to highly complex diagnostic procedures not available in routine laboratories. We show that QuantiFERON assays can be exploited as a simple screening tool that may facilitate adequate and timely treatment. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.
ClusPro PeptiDock: Efficient global docking of peptide recognition motifs using FFT
Apr 21, 2017   Bioinformatics (Oxford, England)
Porter KA, Xia B, Beglov D, Bohnuud T, Alam N, Schueler-Furman O, Kozakov D
ClusPro PeptiDock: Efficient global docking of peptide recognition motifs using FFT
Apr 21, 2017
Bioinformatics (Oxford, England)
We present an approach for the efficient docking of peptide motifs to their free receptor structures. Using a motif based search, we can retrieve structural fragments from the Protein Data Bank (PDB) that are very similar to the peptide's final, bound conformation. We use a Fast Fourier Transform (FFT) based docking method to quickly perform global rigid body docking of these fragments to the receptor. According to CAPRI peptide docking criteria, an acceptable conformation can often be found among the top-ranking predictions. The method is available as part of the protein-protein docking server ClusPro at https://peptidock.cluspro.org/nousername.php . Supplementary data are available at Bioinformatics online.
Dual role of the Toxoplasma gondii clathrin adaptor AP1 in the sorting of rhoptry and microneme proteins and in parasite division
Apr 21, 2017   PLoS Pathogens
Venugopal K, Werkmeister E, Barois N, Saliou JM, Poncet A, Huot L, Sindikubwabo F, Hakimi MA, Langsley G, Lafont F, Marion S
Dual role of the Toxoplasma gondii clathrin adaptor AP1 in the sorting of rhoptry and microneme proteins and in parasite division
Apr 21, 2017
PLoS Pathogens
Toxoplasma gondii possesses a highly polarized secretory system, which efficiently assembles de novo micronemes and rhoptries during parasite replication. These apical secretory organelles release their contents into host cells promoting parasite invasion and survival. Using a CreLox-based inducible knock-out strategy and the ddFKBP over-expression system, we unraveled novel functions of the clathrin adaptor complex TgAP1. First, our data indicate that AP1 in T. gondii likely functions as a conserved heterotetrameric complex composed of the four subunits γ, β, μ1, σ1 and interacts with known regulators of clathrin-mediated vesicular budding such as the unique ENTH-domain containing protein, which we named Epsin-like protein (TgEpsL). Disruption of the μ1 subunit resulted in the mis-sorting of microneme proteins at the level of the Trans-Golgi-Network (TGN). Furthermore, we demonstrated that TgAP1 regulates rhoptry biogenesis by activating rhoptry protein exit from the TGN, but also participates in the post-Golgi maturation process of preROP compartments into apically anchored club-shaped mature organelles. For this latter activity, our data indicate a specific functional relationship between TgAP1 and the Rab5A-positive endosome-like compartment. In addition, we unraveled an original role for TgAP1 in the regulation of parasite division. APμ1-depleted parasites undergo normal daughter cell budding and basal complex assembly but fail to segregate at the end of cytokinesis.
Vaccination strategies against Zika virus
Apr 22, 2017   Current Opinion In Virology
Fernandez E, Diamond MS
Vaccination strategies against Zika virus
Apr 22, 2017
Current Opinion In Virology
The epidemic emergence of Zika virus (ZIKV) in 2015-2016 has been associated with congenital malformations and neurological sequela. Current efforts to develop a ZIKV vaccine build on technologies that successfully reduced infection or disease burden against closely related flaviviruses or other RNA viruses. Subunit-based (DNA plasmid and modified mRNA), viral vectored (adeno- and measles viruses) and inactivated viral vaccines are already advancing to clinical trials in humans after successful mouse and non-human primate studies. Among the greatest challenges for the rapid implementation of immunogenic and protective ZIKV vaccines will be addressing the potential for exacerbating Dengue virus infection or causing Guillain-Barré syndrome through production of cross-reactive immunity targeting related viral or host proteins. Here, we review vaccine strategies under development for ZIKV and the issues surrounding their usage. Copyright © 2017 Elsevier B.V. All rights reserved.

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