bluuebunny/crossref_metadata_2025 huggingface dataset:
Semaphorin4D induces inhibitory synapse formation by rapid stabilization of presynaptic boutons via MET co-activation
Authors: Frias Cátia P. , Bresser Tom , Scheefhals Lisa , Yin Hu Hai , van Bergen en Henegouwen Paul M. P. , Hoogenraad Casper C. , Wierenga Corette J.
Published: January 2017

Changes in inhibitory connections are essential for experience-dependent circuit adaptations. Defects in inhibitory synapses are linked to neurodevelopmental disorders, but the molecular processes underlying inhibitory synapse formation are not well understood. Here we use high resolution two-photon microscopy in organotypic hippocampal slices to examine the signaling pathways induced by the postsynaptic signaling molecule Semaphorin4D (Sema4D) during inhibitory synapse formation. By monitoring changes in individual GFP-labeled presynaptic boutons we found that the primary action of Sema4D is to induce stabilization of presynaptic boutons within tens of minutes. Stabilizing boutons rapidly recruited synaptic vesicles, which was followed by accumulation of postsynaptic gephyrin. Newly formed inhibitory synapses were complete and functional after 24 hours, as determined by electrophysiology and immunohistochemistry. We further showed that Sema4D signaling is regulated by network activity and can induce a local increase in bouton density, suggesting a possible role in circuit adaptation. We further examined the intracellular signaling cascade triggered by Sema4D and found that bouton stabilization occurred through rapid remodeling of actin, and this could be mimicked by the actin-depolymerizing drug Latrunculin B or by reducing ROCK activity. The intracellular signaling cascade required activation of the receptor tyrosine kinase MET, which is a well-known autism risk factor. Our immunohistochemistry data suggests that MET may be localized to presynaptic inhibitory axons. Together, our data yield important insights in the molecular pathway underlying activity-dependent Sema4D-induced synapse formation and reveal a novel role for MET in inhibitory synapses.

Significance Statement

GABAergic synapses provide the main inhibitory control of neuronal activity in the brain. We make important steps in unraveling the molecular processes that take place when formation of inhibitory synapses is triggered by a specific signaling molecule, Sema4D. We find that this process depends on network activity and involves specific remodeling of the intracellular actin cytoskeleton. We also reveal a previously unknown role for MET in inhibitory synapses. As defects in GABAergic synapses have been implied in many brain disorders, and mutations in MET are strong risk factors for autism, our findings urge for a further investigation of the role of MET at inhibitory synapses.
A Network Integration Approach for Drug-Target Interaction Prediction and Computational Drug Repositioning from Heterogeneous Information
Authors: Luo Yunan , Zhao Xinbin , Zhou Jingtian , Yang Jinglin , Zhang Yanqing , Kuang Wenhua , Peng Jian , Chen Ligong , Zeng Jianyang
Published: January 2017

The emergence of large-scale genomic, chemical and pharmacological data provides new opportunities for drug discovery and repositioning. Systematic integration of these heterogeneous data not only serves as a promising tool for identifying new drug-target interactions (DTIs), which is an important step in drug development, but also provides a more complete understanding of the molecular mechanisms of drug action. In this work, we integrate diverse drug-related information, including drugs, proteins, diseases and side-effects, together with their interactions, associations or similarities, to construct a heterogeneous network with 12,015 nodes and 1,895,445 edges. We then develop a new computational pipeline, called DTINet, to predict novel drug-target interactions from the constructed heterogeneous network. Specifically, DTINet focuses on learning a low-dimensional vector representation of features for each node, which accurately explains the topological properties of individual nodes in the heterogeneous network, and then predicts the likelihood of a new DTI based on these representations via a vector space projection scheme. DTINet achieves substantial performance improvement over other state-of-the-art methods for DTI prediction. Moreover, we have experimentally validated the novel interactions between three drugs and the cyclooxygenase (COX) protein family predicted by DTINet, and demonstrated the new potential applications of these identified COX inhibitors in preventing inflammatory diseases. These results indicate that DTINet can provide a practically useful tool for integrating heterogeneous information to predict new drug-target interactions and repurpose existing drugs. The source code of DTINet and the input heterogeneous network data can be downloaded fromhttp://github.com/luoyunan/DTINet.
Single-cell transcriptome analysis of fish immune cells provides insight into the evolution of vertebrate immune cell types
Authors: Carmona Santiago J. , Teichmann Sarah A. , Ferreira Lauren , Macaulay Iain C. , Stubbington Michael J.T. , Cvejic Ana , Gfeller David
Published: January 2017

The immune system of vertebrate species consists of many different cell types that have distinct functional roles and are subject to different evolutionary pressures. Here, we first analyzed conservation of genes specific for all major immune cell types in human and mouse. Our results revealed higher gene turnover and faster evolution of trans-membrane proteins in NK cells compared with other immune cell types, and especially T cells, but similar conservation of nuclear and cytoplasmic protein coding genes. To validate these findings in a distant vertebrate species, we used single-cell RNA sequencing of lck:GFP cells in zebrafish and obtained the first transcriptome of specific immune cell types in a nonmammalian species. Unsupervised clustering and single-cell TCR locus reconstruction identified three cell populations, T cells, a novel type of NK-like cells, and a smaller population of myeloid-like cells. Differential expression analysis uncovered new immune-cell–specific genes, including novel immunoglobulin-like receptors, and neofunctionalization of recently duplicated paralogs. Evolutionary analyses confirmed the higher gene turnover of trans-membrane proteins in NK cells compared with T cells in fish species, suggesting that this is a general property of immune cell types across all vertebrates.
Novel determinants of mammalian primary microRNA processing revealed by systematic evaluation of hairpin-containing transcripts and human genetic variation
Authors: Roden Christine , Gaillard Jonathan , Kanoria Shaveta , Rennie William , Barish Syndi , Cheng Jijun , Pan Wen , Liu Jun , Cotsapas Chris , Ding Ye , Lu Jun
Published: January 2017

Mature microRNAs (miRNAs) are processed from hairpin-containing primary miRNAs (pri-miRNAs). However, rules that distinguish pri-miRNAs from other hairpin-containing transcripts in the genome are incompletely understood. By developing a computational pipeline to systematically evaluate 30 structural and sequence features of mammalian RNA hairpins, we report several new rules that are preferentially utilized in miRNA hairpins and govern efficient pri-miRNA processing. We propose that a hairpin stem length of 36 ± 3 nt is optimal for pri-miRNA processing. We identify two bulge-depleted regions on the miRNA stem, located ∼16–21 nt and ∼28–32 nt from the base of the stem, that are less tolerant of unpaired bases. We further show that the CNNC primary sequence motif selectively enhances the processing of optimal-length hairpins. We predict that a small but significant fraction of human single-nucleotide polymorphisms (SNPs) alter pri-miRNA processing, and confirm several predictions experimentally including a disease-causing mutation. Our study enhances the rules governing mammalian pri-miRNA processing and suggests a diverse impact of human genetic variation on miRNA biogenesis.
Barriers to access to opioid medicines for patients with opioid dependence: a review of legislation and regulations in eleven central and eastern European countries
Authors: Vranken Marjolein J. M. , Mantel‐Teeuwisse Aukje K. , Jünger Saskia , Radbruch Lukas , Scholten Willem , Lisman John A. , Subataite Marija , Schutjens Marie‐Hélène D. B.
Published: January 2017

Background and aims

Barriers linked to drug control systems are considered to contribute to inequitable access to controlled medicines, leaving millions of people in pain and suffering. Most studies focus on access to opioids for the treatment of severe (cancer) pain. This study aims to identify specific access barriers for patients with opioid dependence in legislation and regulations of 11 central and eastern European countries.

Methods

This study builds on a previous analysis of legislation and regulations as part of the EU 7th Framework Access To Opioid Medication in Europe (ATOME) project. An in‐depth analysis was undertaken to determine specific barriers for patients with opioid dependence in need of opioid analgesics or opioid agonist therapy (OAT). For each country, the number and nature of specific potential barriers for these patients were assessed according to eight categories: prescribing; dispensing; manufacturing; usage; trade and distribution; affordability; penalties; and other. An additional keyword search was conducted to minimize the omission of barriers. Barriers in an additional category, language, were recorded qualitatively. Countries included Bulgaria, Cyprus, Estonia, Greece, Hungary, Latvia, Lithuania, Serbia, Slovakia, Slovenia and Turkey.

Results

Ten of the 11 countries (all except Estonia) showed specific potential barriers in their legislation and regulations. The total number of barriers varied from two (Slovenia) to 46 (Lithuania); the number of categories varied from one (Slovenia) to five (Lithuania). Most specific potential barriers were shown in the categories ‘prescribing’, ‘usage’ and ‘other’. The total number in a single category varied from one to 18 (Lithuania, prescribing). Individual differences between countries in the same specific potential barrier were shown; for example, variation in minimum age criteria for admission to OAT ranging from 15 (Lithuania, in special cases) to 20 years (Greece). All countries had stigmatizing language in their legislation.

Conclusions

Patients with opioid dependence are likely to experience specific barriers to accessing opioids in addition to those experienced by other non‐dependent patients.
Migration as an Adaptation Strategy for Atoll Island States
Authors: Yamamoto Lilian , Esteban Miguel
Published: January 2017

Adopting a policy of migration can be one possible adaptation strategy against climate change. It has been forecasted that if the worst predictions regarding climate change and sea level rise become reality atolls around the world could become submerged in the future. This would render them uninhabitable and could lead to questions about whether Atoll Island States could still be considered as States. The international community has been avoiding any commitment to create a convention that would protect people displaced by climate change. In order to solve such potential problems, the authors will argue that a framework of bilateral agreements, initiatives, and national policies could constitute a viable solution for the various interested parties. The article will discuss the characteristics of Atoll Island States, touching on possible solutions for climate change displacement which have been discussed by the governments and civil society of the affected States.
Integration by Stealth: How the European Union Gained Competence over Foreign Direct Investment
Authors: Meunier Sophie
Published: January 2017

How are policy competences allocated between different actors? This article contributes to the literature on institutional development through an in‐depth case‐study of the conditions under which the competence over the negotiation of agreements on foreign direct investment (FDI) was transferred from the national level to the European Union (EU) in the 2009 Lisbon Treaty. Most analysts assume that this competence shift was a rationally designed delegation, intended to maximize European bargaining power in international investment negotiations and conceived as an important element of a teleological drive to make the EU a meaningful external actor. This article tells a different story – one where the competence shift happened by stealth as a result of a combination of neo‐functionalist Commission entrepreneurship and historical accident, against the preferences of the Member States. The article also assesses whether the conditions under which the competence was transferred have implications for the implementation of the new policy.
Prophylactic tributyrin treatment mitigates chronic‐binge ethanol‐induced intestinal barrier and liver injury
Authors: Cresci Gail A , Glueck Bryan , McMullen Megan R , Xin Wei , Allende Daniella , Nagy Laura E
Published: January 2017

Background and Aim

Impaired gut‐liver axis is a potential factor contributing to alcoholic liver disease. Ethanol depletes intestinal integrity and causes gut dysbiosis. Butyrate, a fermentation byproduct of gut microbiota, is altered negatively following chronic ethanol exposure. This study aimed to determine whether prophylactic tributyrin could protect the intestinal barrier and liver in mice during combined chronic‐binge ethanol exposure.

Methods

C57BL/6J mice exposed to 5% v/v ethanol‐containing diet for 10 days received a single ethanol gavage (5 g/kg) 9 h before euthanasia. Control mice were isocalorically pair‐fed maltose dextrin for ethanol. Diets were supplemented (5 mM) with tributyrin or glycerol. Intestine and liver disease activity was assessed histologically. Protein and mRNA expression of tight junction (TJ) proteins, toll‐like receptors, and tumor necrosis factor‐alpha were assessed. Caco‐2 monolayers with or without ethanol exposure and/or sodium butyrate were used to test butyrate's direct effects on intestinal integrity.

Results

Chronic‐binge ethanol feeding impaired intestinal TJ protein co‐localization staining; however, tributyrin co‐treatment mitigated these effects. Ethanol depleted TJ and transepithelial electrical resistance in Caco‐2 monolayers, but butyrate co‐treatment reduced these effects. Hepatic toll‐like receptor mRNA expression and tumor necrosis factor‐alpha protein expression was induced by ethanol; however, the response was significantly dampened in mice co‐treated with tributyrin. Tributyrin altered localization of both neutrophils and single hepatocyte death: Leukocytes and apoptotic hepatocytes localized predominantly around the portal tract in ethanol‐only treated mice, whereas localization predominated around the central vein in ethanol‐tributyrin mice.

Conclusions

Prophylactic tributyrin supplementation mitigated effects of combined chronic‐binge ethanol exposure on disruption of intestinal TJ localization and intestinal permeability and liver injury.
Association of granulomas in mesenteric lymph nodes in Crohn's disease with younger age and transmural inflammation
Authors: Sun Xingwen , Yuan Lisi , Li Yi , Shen Bo , Xie Hao , Liu Xiuli
Published: January 2017

Background and Aims

Mesenteric lymph nodes were usually removed during bowel resection surgery for Crohn's disease (CD). Mesenteric lymph node (MLN) granuloma predicted postoperative disease recurrence in patients with CD after ileocolic resection (ICR). This study was aimed to identify factors associated with MLN granulomas in a cohort of CD patients who underwent ICR.

Methods

The study group consisted of 18 patients with CD who underwent the index ICR between 2004 and 2012 and had MLN granulomas. The control group included 32 cases negative for MLN granulomas, which were randomly selected from the same period. Clinicodemographic and histological features were compared in these two groups.

Results

The presence of MLN granuloma was associated with younger age, perianal disease, and ileocolic disease. Gender, body mass index, smoking status, disease duration, extraintestinal manifestation, length of bowel resected, total number of nodes examined, medical treatment, and indications for surgery were not significantly different between the groups. The presence of MLN granuloma was associated with mural granuloma, transmural inflammation, mural abscess, hypertrophy of the muscularis propria, fibrosis of the muscularis propria, and serosal fibrosis in the small bowel. The multivariable analysis confirms that MLN granuloma was associated with age and transmural inflammation.

Conclusions

In CD patients who underwent the index ICR, the presence of MLN granuloma was associated with younger age and transmural inflammation. Our current results suggest that the detection of MLN granulomas may be enhanced in patients at higher risk, that is, younger patients or those with transmural inflammation on imaging or histopathology.
Established and emerging methods for assessment of small and large intestinal motility
Authors: Grønlund D. , Poulsen J. L. , Sandberg T. H. , Olesen A. E. , Madzak A. , Krogh K. , Frøkjær J. B. , Drewes A. M.
Published: January 2017

Background

Gastrointestinal symptoms are common in the general population and may originate from disturbances in gut motility. However, fundamental mechanistic understanding of motility remains inadequate, especially of the less accessible regions of the small bowel and colon. Hence, refinement and validation of objective methods to evaluate motility of the whole gut is important. Such techniques may be applied in clinical settings as diagnostic tools, in research to elucidate underlying mechanisms of diseases, and to evaluate how the gut responds to various drugs. A wide array of such methods exists; however, a limited number are used universally due to drawbacks like radiation exposure, lack of standardization, and difficulties interpreting data. In recent years, several new methods such as the 3D‐Transit system and magnetic resonance imaging assessments on small bowel and colonic motility have emerged, with the advantages that they are less invasive, use no radiation, and provide much more detailed information.

Purpose

This review outlines well‐established and emerging methods to evaluate small bowel and colonic motility in clinical settings and in research. The latter include the 3D‐Transit system, magnetic resonance imaging assessments, and high‐resolution manometry. Procedures, indications, and the relative strengths and weaknesses of each method are summarized.
Gastrointestinal disorders in joint hypermobility syndrome/Ehlers‐Danlos syndrome hypermobility type: A review for the gastroenterologist
Authors: Beckers A. B. , Keszthelyi D. , Fikree A. , Vork L. , Masclee A. , Farmer A. D. , Aziz Q.
Published: January 2017

Background

Joint hypermobility syndrome (JHS)/Ehlers‐Danlos syndrome hypermobility type (EDS‐HT) is the most common hereditary non‐inflammatory disorder of connective tissue, characterized by a wide range of symptoms, mainly joint hyperextensibility and musculoskeletal symptoms. A majority of patients also experiences gastrointestinal (GI) symptoms. Furthermore, JHS/EDS‐HT has specifically been shown to be highly prevalent in patients with functional GI disorders, such as functional dyspepsia and irritable bowel syndrome.

Purpose

The aim of this review was to examine the nature of GI symptoms and their underlying pathophysiology in JHS/EDS‐HT. In addition, we consider the clinical implications of the diagnosis and treatment of JHS/EDS‐HT for practicing clinicians in gastroenterology. Observations summarized in this review may furthermore represent the first step toward the identification of a new pathophysiological basis for a substantial subgroup of patients with functional GI disorders.
ON THE BUREAUCRACY OF BUREAUCRACIES: ANALYSING THE SIZE AND ORGANIZATION OF OVERHEAD IN PUBLIC ORGANIZATIONS
Authors: BOON JAN , WYNEN JAN
Published: January 2017

Governments across the globe try to rebalance their budgets by rationalizing overhead operations. When overhead‐reducing policies are adopted, it is important to understand why some central government organizations have a higher overhead than others, and why organizational models to produce overhead efficiencies are used to different degrees. This study focuses on the Flemish context to analyse differences between central government organizations in the size and organization of two overhead processes: human resources management (HRM) and finance and control (FIN). Significant effects are found for autonomy, organizational size, spatial dispersion and budgetary stress, yet effects vary according to whether HRM or FIN is considered and whether the focus is on the size or the organization of HRM or FIN. Our findings have practical implications to get a process‐sensitive understanding of the size and organization of overhead, and theoretical implications as they cast light on factors that shape decision‐making in public organizations.
The Formation of Bridging and Bonding Social Capital to Empower Mothers from Low-income Families
Authors: Chawa Anif Fatma
Published: January 2017

This article presents empirical findings of research seeking to explore the formation of social capital in community development practice, that exhibits a model or mechanism by which social capital can be formed and maintained. This study also depicts how the empowerment objective can be achieved through this form of social capital. The result of this study found that the grouping mechanism established by local community development organization, namely Posdaya, has led to the formation and strengthening of social capital. The members of Posdaya have been grouped based the similarities of their interests, such as hobbies, talents, and professions. This study highlights the fact that this social capital has given rise to collective participation of the Posdaya members by sharing their knowledge, skills, and information; and finding solutions to their development problems with minimum outside intervention. These mutual and collective actions can be considered part of the self-help principle which has succeeded in empowering the members of Posdaya.
The Emergence and Development of National Congress Party in Sudan (1998-2005)
Authors: Jamie Faiz Omar Mohammad
Published: January 2017

The paper analyses the emergence of the National Congress Party (N.C.P.) within the context of the recent political history of Sudan in post 1989 era. The date marks the time when Islamists in Sudan assumed power following a coup d’état led by General Omer Al-Bashir, latter on came to be known as the Ingaz regime. The significance of the experience of this Party emanates from the fact that, though it started as a one ruling party, it managed to conclude in 2005 the Comprehensive Peace Agreement (C.P.A.) a major peace agreement ending one of the longest internal wars in Africa. The Agreement was included in the Constitution of the country, consequently hosting Sudan People’s Liberation Movement (SPLM), the movement which led the rebellion to become a partner party/movement in the rule of the country. The paper reflects on this power sharing experience during the Interim Period up to 2011 wherein the Referendum on Self-determination of South Sudan resulted in dividing the Country into two sovereign states.
Simulated microgravity decreases circulating iron in rats: role of inflammation‐induced hepcidin upregulation
Authors: Cavey Thibault , Pierre Nicolas , Nay Kévin , Allain Coralie , Ropert Martine , Loréal Olivier , Derbré Frédéric
Published: January 2017

New Findings

What is the central question of this study?

Although microgravity is well known to reduce circulating iron in astronauts, the underlying mechanism is still unknown. We investigated whether hepcidin, a key hormone regulating iron metabolism, could be involved in this deleterious effect.

What is the main finding and its importance?

We show that hindlimb suspension, a model of microgravity, stimulates the production of hepcidin in liver of rats. In agreement with the biological role of hepcidin, we found a decrease of circulating iron and an increase of spleen iron content in hindlimb‐unloaded rats. Consequently, our study supports the idea that hepcidin could play a role in the alteration of iron metabolism parameters observed during spaceflight.

During spaceflight, humans exposed to microgravity exhibit an increase of iron storage and a reduction of circulating iron. Such perturbations could promote oxidative stress and anaemia in astronauts. The mechanism by which microgravity modulates iron metabolism is still unknown. Herein, we hypothesized that microgravity upregulates hepcidin, a hormone produced by the liver that is the main controller of iron homeostasis. To test this hypothesis, rats were submitted to hindlimb unloading (HU), the reference model to mimic the effects of microgravity in rodents. After 7 days, the mRNA level of hepcidin was increased in the liver of HU rats (+74%, P = 0.001). In agreement with the biological role of hepcidin, we found an increase of spleen iron content (+78%, P = 0.030) and a decrease of serum iron concentration (−35%, P = 0.002) and transferrin saturation (−25%, P = 0.011) in HU rats. These findings support a role of hepcidin in microgravity‐induced iron metabolism alteration. Furthermore, among the signalling pathways inducing hepcidin mRNA expression, we found that only the interleukin‐6/signal transducer and activator of transcription 3 (IL‐6/STAT3) axis was activated by HU, as shown by the increase of phospho‐STAT3 (+193%, P < 0.001) and of the hepatic mRNA level of haptoglobin (+167%, P < 0.001), a STAT3‐inducible gene, in HU rats. Taken together, these data support the idea that microgravity may alter iron metabolism through an inflammatory process upregulating hepcidin.
Role of STIM2 in cell function and physiopathology
Authors: Berna‐Erro Alejandro , Jardin Isaac , Salido Gines M. , Rosado Juan A.
Published: January 2017

An endoplasmic reticulum (ER)‐resident protein that regulates cytosolic and ER free‐Ca2+ concentration by induction of store‐operated calcium entry: that is the original definition of STIM2 and its function. While its activity strongly depends on the amount of calcium stored in the ER, its function goes further, to intracellular signalling and gene expression. Initially under‐studied owing to the prominent function of STIM1, STIM2 came to be regarded as vital in mice, gradually emerging as an important player in the nervous system, and cooperating with STIM1 in the immune system. STIM2 has also been proposed as a relevant player in pathological conditions related to ageing, Alzheimer's and Huntington's diseases, autoimmune disorders and cancer. The discovery of additional functions, together with new splicing forms with opposite roles, has clarified existing controversies about STIM2 function in SOCE. With STIM2 being essential for life, but apparently not for development, newly available data demonstrate a complex and still intriguing behaviour that this review summarizes, updating current knowledge of STIM2 function.

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Listening to each other: Infectious disease and cancer immunology
Authors: Vance Russell E. , Eichberg Michael J. , Portnoy Daniel A. , Raulet David H.
Published: January 2017

Shared immunological principles influence design of immunotherapies for both cancer and infectious diseases.
Efficient Low-pH Iron Removal by a Microbial Iron Oxide Mound Ecosystem at Scalp Level Run
Authors: Grettenberger Christen L. , Pearce Alexandra R. , Bibby Kyle J. , Jones Daniel S. , Burgos William D. , Macalady Jennifer L.
Published: January 2017

Acid mine drainage (AMD) is a major environmental problem affecting tens of thousands of kilometers of waterways worldwide. Passive bioremediation of AMD relies on microbial communities to oxidize and remove iron from the system; however, iron oxidation rates in AMD environments are highly variable among sites. At Scalp Level Run (Cambria County, PA), first-order iron oxidation rates are 10 times greater than at other coal-associated iron mounds in the Appalachians. We examined the bacterial community at Scalp Level Run to determine whether a unique community is responsible for the rapid iron oxidation rate. Despite strong geochemical gradients, including a >10-fold change in the concentration of ferrous iron from 57.3 mg/liter at the emergence to 2.5 mg/liter at the base of the coal tailings pile, the bacterial community composition was nearly constant with distance from the spring outflow. Scalp Level Run contains many of the same taxa present in other AMD sites, but the community is dominated by two strains of
Ferrovum myxofaciens
, a species that is associated with high rates of Fe(II) oxidation in laboratory studies.

IMPORTANCE
Acid mine drainage pollutes more than 19,300 km of rivers and streams and 72,000 ha of lakes worldwide. Remediation is frequently ineffective and costly, upwards of $100 billion globally and nearly $5 billion in Pennsylvania alone. Microbial Fe(II) oxidation is more efficient than abiotic Fe(II) oxidation at low pH (P. C. Singer and W. Stumm, Science 167:1121–1123, 1970,
https://doi.org/10.1126/science.167.3921.1121
). Therefore, AMD bioremediation could harness microbial Fe(II) oxidation to fuel more-cost-effective treatments. Advances will require a deeper understanding of the ecology of Fe(II)-oxidizing microbial communities and the factors that control their distribution and rates of Fe(II) oxidation. We investigated bacterial communities that inhabit an AMD site with rapid Fe(II) oxidation and found that they were dominated by two operational taxonomic units (OTUs) of
Ferrovum myxofaciens
, a taxon associated with high laboratory rates of iron oxidation. This research represents a step forward in identifying taxa that can be used to enhance cost-effective AMD bioremediation.
Engineering Mycobacteria for the Production of Self-Assembling Biopolyesters Displaying Mycobacterial Antigens for Use as a Tuberculosis Vaccine
Authors: Lee Jason W. , Parlane Natalie A. , Rehm Bernd H. A. , Buddle Bryce M. , Heiser Axel
Published: January 2017

Tuberculosis (TB) is a disease caused by
Mycobacterium tuberculosis
or
Mycobacterium bovis
and still remains one of the world's biggest global health burdens. Recently, engineered polyhydroxyalkanoate (PHA) biobeads that were produced in both
Escherichia coli
and
Lactococcus lactis
and displayed mycobacterial antigens were found to induce significant cell-mediated immune responses in mice. We observed that such PHA beads contained host cell proteins as impurities, which we hypothesized to have the potential to induce immunity. In this study, we aimed to develop PHA beads produced in mycobacteria (mycobacterial PHA biobeads [MBB]) and test their potential as a TB vaccine in a mouse model. As a model organism, nonpathogenic
Mycobacterium smegmatis
was engineered to produce MBB or MBB with immobilized mycobacterial antigens Ag85A and ESAT-6 on their surface (A:E-MBB). Three key enzymes involved in the poly(3-hydroxybutyric acid) pathway, namely, β-ketothiolase (PhaA), acetoacetyl-coenzyme A reductase (PhaB), and PHA synthase (PhaC), were engineered into
E. coli
-
Mycobacterium
shuttle plasmids and expressed in
trans
. Immobilization of specific antigens to the surface of the MBB was achieved by creating a fusion with the PHA synthase which remains covalently attached to the polyester core, resulting in PHA biobeads displaying covalently immobilized antigens. MBB, A:E-MBB, and an
M. smegmatis
vector control (MVC) were used in a mouse immunology trial, with comparison to phosphate-buffered saline (PBS)-vaccinated and
Mycobacterium bovis
BCG-vaccinated groups. We successfully produced MBB and A:E-MBB and used them as vaccines to induce a cellular immune response to mycobacterial antigens.

IMPORTANCE
Tuberculosis (TB) is a disease caused by
Mycobacterium tuberculosis
or
Mycobacterium bovis
and still remains one of the world's biggest global health burdens. In this study, we produced polyhydroxyalkanoate (PHA) biobeads in mycobacteria and used them as vaccines to induce a cellular immune response to mycobacterial antigens.
Establishment and Validation of RNA-Based Predictive Models for Understanding Survival of Vibrio parahaemolyticus in Oysters Stored at Low Temperatures
Authors: Liao Chao , Zhao Yong , Wang Luxin
Published: January 2017

This study developed RNA-based predictive models describing the survival of
Vibrio parahaemolyticus
in Eastern oysters (
Crassostrea virginica
) during storage at 0, 4, and 10°C. Postharvested oysters were inoculated with a cocktail of five
V. parahaemolyticus
strains and were then stored at 0, 4, and 10°C for 21 or 11 days. A real-time reverse transcription-PCR (RT-PCR) assay targeting expression of the
tlh
gene was used to evaluate the number of surviving
V. parahaemolyticus
cells, which was then used to establish primary molecular models (MMs). Before construction of the MMs, consistent expression levels of the
tlh
gene at 0, 4, and 10°C were confirmed, and this gene was used to monitor the survival of the total
V. parahaemolyticus
cells. In addition, the
tdh
and
trh
genes were used for monitoring the survival of virulent
V. parahaemolyticus
. Traditional models (TMs) were built based on data collected using a plate counting method. From the MMs,
V. parahaemolyticus
populations had decreased 0.493, 0.362, and 0.238 log
10
CFU/g by the end of storage at 0, 4, and 10°C, respectively. Rates of reduction of
V. parahaemolyticus
shown in the TMs were 2.109, 1.579, and 0.894 log
10
CFU/g for storage at 0, 4, and 10°C, respectively. Bacterial inactivation rates (IRs) estimated with the TMs (−0.245, −0.152, and −0.121 log
10
CFU/day, respectively) were higher than those estimated with the MMs (−0.134, −0.0887, and −0.0732 log
10
CFU/day, respectively) for storage at 0, 4, and 10°C. Higher viable
V. parahaemolyticus
numbers were predicted using the MMs than using the TMs. On the basis of this study, RNA-based predictive MMs are the more accurate and reliable models and can prevent false-negative results compared to TMs.

IMPORTANCE
One important method for validating postharvest techniques and for monitoring the behavior of
V. parahaemolyticus
is to establish predictive models. Unfortunately, previous predictive models established based on plate counting methods or on DNA-based PCR can underestimate or overestimate the number of surviving cells. This study developed and validated RNA-based molecular predictive models to describe the survival of
V. parahaemolyticus
in oysters during low-temperature storage (0, 4, and 10°C). The RNA-based predictive models show the advantage of being able to count all of the culturable, nonculturable, and stressed cells. By using primers targeting the
tlh
gene and pathogenesis-associated genes (
tdh
and
trh
), real-time RT-PCR can evaluate the total surviving
V. parahaemolyticus
population as well as differentiate the pathogenic ones from the total population. Reliable and accurate predictive models are very important for conducting risk assessment and management of pathogens in food.
Rapid Enrichment of Dehalococcoides-Like Bacteria by Partial Hydrophobic Separation
Authors: Temme Hanna R. , Sande Kipp , Yan Tao , Novak Paige J.
Published: January 2017

Organohalide-respiring bacteria can be difficult to enrich and isolate, which can limit research on these important organisms. The goal of this research was to develop a method to rapidly (minutes to days) enrich these organisms from a mixed community. The method presented is based on the hypothesis that organohalide-respiring bacteria would be more hydrophobic than other bacteria as they dehalogenate hydrophobic compounds. The method developed tests this hypothesis by separating a portion of putative organohalide-respiring bacteria, those phylogenetically related to
Dehalococcoides mccartyi
, at the interface between a hydrophobic organic solvent and an aqueous medium. This novel partial separation technique was tested with a polychlorinated biphenyl-enriched sediment-free culture, a tetrachloroethene-enriched digester sludge culture, and uncontaminated lake sediment. Significantly higher fractions, up to 20.4 times higher, of putative organohalide-respiring bacteria were enriched at the interface between the medium and either hexadecane or trichloroethene. The selective partial separation of these putative organohalide-respiring bacteria occurred after 20 min, strongly suggesting that the separation was a result of physical-chemical interactions between the cell surface and hydrophobic solvent. Dechlorination activity postseparation was verified by the production of
cis
-dichloroethene when amended with tetrachloroethene. A longer incubation time of 6 days prior to separation with trichloroethene increased the total number of putative organohalide-respiring bacteria. This method provides a way to quickly separate some of the putative organohalide-respiring bacteria from other bacteria, thereby improving our ability to study multiple and different bacteria of potential interest and improving knowledge of these bacteria.

IMPORTANCE
Organohalide-respiring bacteria, bacteria capable of respiring chlorinated contaminants, can be difficult to enrich, which can limit their predictable use for the bioremediation of contaminated sites. This paper describes a method to quickly separate
Dehalococcoides
-like bacteria, a group of organisms containing organohalide-respiring bacteria, from other bacteria in a mixed community. From this work,
Dehalococcoides
-like bacteria appear to have a hydrophobic cell surface, facilitating a rapid (20 min) partial separation from a mixed culture at the surface of a hydrophobic liquid. This method was verified in a polychlorinated biphenyl-enriched sediment-free culture, an anaerobic digester sludge, and uncontaminated sediment. The method described can drastically reduce the amount of time required to partially separate
Dehalococcoides
-like bacteria from a complex mixed culture, improving researchers' ability to study these important bacteria.
Polysaccharide Degradation Capability of Actinomycetales Soil Isolates from a Semiarid Grassland of the Colorado Plateau
Authors: Yeager Chris M. , Gallegos-Graves La Verne , Dunbar John , Hesse Cedar N. , Daligault Hajnalka , Kuske Cheryl R.
Published: January 2017

Among the bacteria, members of the order
Actinomycetales
are considered quintessential degraders of complex polysaccharides in soils. However, studies examining complex polysaccharide degradation by
Actinomycetales
(other than
Streptomyces
spp.) in soils are limited. Here, we examine the lignocellulolytic and chitinolytic potential of 112
Actinomycetales
strains, encompassing 13 families, isolated from a semiarid grassland of the Colorado Plateau in Utah. Members of the
Streptomycetaceae
,
Pseudonocardiaceae
,
Micromonosporaceae
, and
Promicromonosporaceae
families exhibited robust activity against carboxymethyl cellulose, xylan, chitin, and pectin substrates (except for low/no pectinase activity by the
Micromonosporaceae
). When incubated in a hydrated mixture of blended
Stipa
and
Hilaria
grass biomass over a 5-week period,
Streptomyces
and
Saccharothrix
(a member of the
Pseudonocardiaceae
) isolates produced high levels of extracellular enzyme activity, such as endo- and exocellulase, glucosidase, endo- and exoxylosidase, and arabinofuranosidase. These characteristics make them well suited to degrade the cellulose and hemicellulose components of grass cell walls. On the basis of the polysaccharide degradation profiles of the isolates, relative abundance of
Actinomycetales
sequences in 16S rRNA gene surveys of Colorado Plateau soils, and analysis of genes coding for polysaccharide-degrading enzymes among 237
Actinomycetales
genomes in the CAZy database and 5 genomes from our isolates, we posit that
Streptomyces
spp. and select members of the
Pseudonocardiaceae
and
Micromonosporaceae
likely play an important role in the degradation of hemicellulose, cellulose, and chitin substances in dryland soils.

IMPORTANCE
Shifts in the relative abundance of
Actinomycetales
taxa have been observed in soil microbial community surveys during large, manipulated climate change field studies. However, our limited understanding of the ecophysiology of diverse
Actinomycetales
taxa in soil systems undermines attempts to determine the underlying causes of the population shifts or their impact on carbon cycling in soil. This study combines a systematic analysis of the polysaccharide degradation potential of a diverse collection of
Actinomycetales
isolates from surface soils of a semiarid grassland with analysis of genomes from five of these isolates and publicly available
Actinomycetales
genomes for genes encoding polysaccharide-active enzymes. The results address an important gap in knowledge of
Actinomycetales
ecophysiology—identification of key taxa capable of facilitating lignocellulose degradation in dryland soils. Information from this study will benefit future metagenomic studies related to carbon cycling in dryland soils by providing a baseline linkage of
Actinomycetales
phylogeny with lignocellulolytic functional potential.
Lysobacter PilR, the Regulator of Type IV Pilus Synthesis, Controls Antifungal Antibiotic Production via a Cyclic di-GMP Pathway
Authors: Chen Yuan , Xia Jing , Su Zhenhe , Xu Gaoge , Gomelsky Mark , Qian Guoliang , Liu Fengquan
Published: January 2017

Lysobacter enzymogenes
is a ubiquitous soil gammaproteobacterium that produces a broad-spectrum antifungal antibiotic, known as heat-stable antifungal factor (HSAF). To increase HSAF production for use against fungal crop diseases, it is important to understand how HSAF synthesis is regulated. To gain insights into transcriptional regulation of the HSAF synthesis gene cluster, we generated a library with deletion mutations in the genes predicted to encode response regulators of the two-component signaling systems in
L. enzymogenes
strain OH11. By quantifying HSAF production levels in the 45 constructed mutants, we identified two strains that produced significantly smaller amounts of HSAF. One of the mutations affected a gene encoding a conserved bacterial response regulator, PilR, which is commonly associated with type IV pilus synthesis. We determined that
L. enzymogenes
PilR regulates pilus synthesis and twitching motility via a traditional pathway, by binding to the
pilA
promoter and upregulating
pilA
expression. Regulation of HSAF production by PilR was found to be independent of pilus formation. We discovered that the
pilR
mutant contained significantly higher intracellular levels of the second messenger cyclic di-GMP (c-di-GMP) and that this was the inhibitory signal for HSAF production. Therefore, the type IV pilus regulator PilR in
L. enzymogenes
activates twitching motility while downregulating antibiotic HSAF production by increasing intracellular c-di-GMP levels. This study identifies a new role of a common pilus regulator in proteobacteria and provides guidance for increasing antifungal antibiotic production in
L. enzymogenes
.

IMPORTANCE
PilR is a widespread response regulator of the two-component system known for regulating type IV pilus synthesis in proteobacteria. Here we report that, in the soil bacterium
Lysobacter enzymogenes
, PilR regulates pilus synthesis and twitching motility, as expected. Unexpectedly, PilR was also found to control intracellular levels of the second messenger c-di-GMP, which in turn inhibits production of the antifungal antibiotic HSAF. The coordinated production of type IV pili and antifungal antibiotics has not been observed previously.
Mechanisms of Horizontal Cell-to-Cell Transfer of Wolbachia spp. in Drosophila melanogaster
Authors: White Pamela M. , Pietri Jose E. , Debec Alain , Russell Shelbi , Patel Bhavin , Sullivan William
Published: January 2017

Wolbachia
is an intracellular endosymbiont present in most arthropod and filarial nematode species. Transmission between hosts is primarily vertical, taking place exclusively through the female germ line, although horizontal transmission has also been documented. The results of several studies indicate that
Wolbachia
spp. can undergo transfer between somatic and germ line cells during nematode development and in adult flies. However, the mechanisms underlying horizontal cell-to-cell transfer remain largely unexplored. Here, we establish a tractable system for probing horizontal transfer of
Wolbachia
cells between
Drosophila melanogaster
cells in culture using fluorescence
in situ
hybridization (FISH). First, we show that horizontal transfer is independent of cell-to-cell contact and can efficiently take place through the culture medium within hours. Further, we demonstrate that efficient transfer utilizes host cell phagocytic and clathrin/dynamin-dependent endocytic machinery. Lastly, we provide evidence that this process is conserved between species, showing that horizontal transfer from mosquito to
Drosophila
cells takes place in a similar fashion. Altogether, our results indicate that
Wolbachia
utilizes host internalization machinery during infection, and this mechanism is conserved across insect species.

IMPORTANCE
Our work has broad implications for the control and treatment of tropical diseases.
Wolbachia
can confer resistance against a variety of human pathogens in mosquito vectors. Elucidating the mechanisms of horizontal transfer will be useful for efforts to more efficiently infect nonnatural insect hosts with
Wolbachia
as a biological control agent. Further, as
Wolbachia
is essential for the survival of filarial nematodes, understanding horizontal transfer might provide new approaches to treating human infections by targeting
Wolbachia
. Finally, this work provides a key first step toward the genetic manipulation of
Wolbachia
.
Use of Acupuncture in the Usa: Changes over a Decade (2002–2012)
Authors: Cui Jia , Wang Shaobai , Ren Jiehui , Zhang Jun , Jing Jun
Published: January 2017

Background

The use of acupuncture has gained popularity in the USA. The number of acupuncture users and licensed acupuncturists increased by 50% and 100%, respectively, between 2002 and 2012, coinciding with increasing acknowledgement of the importance and efficacy of acupuncture over this time period.

Methods

This paper presents new findings from the complementary health approaches section of the 2012 National Health Interview Survey (n=33 373 respondents). In particular, data on the use of acupuncture and user characteristics were compared against data collected from an earlier survey in 2002. Statistical analyses included weighted distribution, logistical regression and Pearson's χ2 tests.

Results

The profile of the most common acupuncture users comprised the following sociodemographics age 41–65 years (47.4%); female gender (69.6%); and non-Hispanic (85.3%) and/or white (78.1%) ethnicity. Respondents also tended to be US citizens (92.1%) with some college education (57.1%) and in very good to excellent health (60.8%). The proportion of respondents using acupuncture for treatment of a specific health problem, as opposed to promotion of general wellness, was 84.7% in 2002 and 55.3% in 2012.

Conclusions

Our data suggest a growing development of acupuncture in the USA. We anticipate that the findings of our analysis of the changes in acupuncture use over the 10-year period from 2002 to 2012 will provide information for users, acupuncturists, researchers and the general public to help better understand the status of acupuncture and reasons for its usage, and to anticipate future trends for acupuncture use in the USA.