bluuebunny/crossref_metadata_2025 huggingface dataset:
Design and synthesis of a minimal bacterial genome
Authors: Hutchison Clyde A. , Chuang Ray-Yuan , Noskov Vladimir N. , Assad-Garcia Nacyra , Deerinck Thomas J. , Ellisman Mark H. , Gill John , Kannan Krishna , Karas Bogumil J. , Ma Li , Pelletier James F. , Qi Zhi-Qing , Richter R. Alexander , Strychalski Elizabeth A. , Sun Lijie , Suzuki Yo , Tsvetanova Billyana , Wise Kim S. , Smith Hamilton O. , Glass John I. , Merryman Chuck , Gibson Daniel G. , Venter J. Craig
Published: March 2016

Designing and building a minimal genome

A goal in biology is to understand the molecular and biological function of every gene in a cell. One way to approach this is to build a minimal genome that includes only the genes essential for life. In 2010, a 1079-kb genome based on the genome of
Mycoplasma mycoides
(JCV-syn1.0) was chemically synthesized and supported cell growth when transplanted into cytoplasm. Hutchison III
et al.
used a design, build, and test cycle to reduce this genome to 531 kb (473 genes). The resulting JCV-syn3.0 retains genes involved in key processes such as transcription and translation, but also contains 149 genes of unknown function.

Science
, this issue p.
10.1126/science.aad6253
Catalytic borylation of methane
Authors: Smith Kyle T. , Berritt Simon , González-Moreiras Mariano , Ahn Seihwan , Smith Milton R. , Baik Mu-Hyun , Mindiola Daniel J.
Published: March 2016

Methane borylation in a cyclohexane sea

Although methane combusts readily at high temperatures, it is generally the hardest hydrocarbon to transform under gentler conditions, owing to its particularly strong C-H bonds. Cook
et al.
now show that soluble rhodium, iridium, and ruthenium catalysts can slice through these C-H bonds to add boron substituents to methane at 150°C. Smith
et al.
report the iridium-catalyzed reaction using phosphine ligands to enhance activity. Both studies were performed in cyclohexane solvent, revealing a remarkable selective preference for the methane reaction over functionalization of the cyclic hydrocarbon.

Science
, this issue pp.
1421
and
1424
The paradigm shift, in perspective

Kuhn's
Structure of Scientific Revolutions
at Fifty Reflections on a Science Classic

Robert J. Richards and Lorraine Daston (Eds.)
University of Chicago Press, 2016. 234 pp.
Authors: Mitchell Sandra D.
Published: March 2016

A new collection of essays takes a fresh look at Thomas Kuhn's classic text
The social gene
The Society of Genes
Itai Yanai and Martin Lercher
Harvard University Press, 2016. 294 pp
Authors: Swift Joseph
Published: March 2016

Advances in genetic research prompt a pair of scientists to update the “selfish gene” metaphor
Pneumococcal Vaccination in High-Risk Individuals: Are We Doing It Right?
Authors: Papadatou Ioanna , Spoulou Vana
Published: March 2016

Controversy exists regarding the optimal use of the 23-valent pneumococcal conjugate vaccine for the protection of high-risk individuals, such as children and adults with immunocompromising conditions and the elderly. The effectiveness and immunogenicity of 23-valent pneumococcal polysaccharide vaccine (PPV23) are limited in such high-risk populations compared to the healthy, with meta-analyses failing to provide robust evidence on vaccine efficacy against invasive pneumococcal disease (IPD) or pneumonia. Moreover, several studies have demonstrated a PPV23-induced state of immune tolerance or hyporesponsiveness to subsequent vaccination, where the response to revaccination does not reach the levels achieved with primary vaccination. The clinical significance of hyporesponsiveness is not yet clarified, but attenuated humoral and cellular response could lead to reduced levels of protection and increased susceptibility to pneumococcal disease. As disease epidemiology among high-risk groups shows that we are still in need of maximum serotype coverage, the optimal use of PPV23 in the context of combined conjugate/polysaccharide vaccine schedules is an important priority. In this minireview, we discuss PPV23-induced hyporesponsiveness and its implications in designing highly effective vaccination schedules for the optimal protection for high-risk individuals.
Performance Characteristics of FilmArray Respiratory Panel v1.7 for Detection of Adenovirus in a Large Cohort of Pediatric Nasopharyngeal Samples: One Test May Not Fit All
Authors: Song Eunkyung , Wang Huanyu , Salamon Doug , Jaggi Preeti , Leber Amy
Published: March 2016

The FilmArray Respiratory Panel (RP) v1.7 assay has improved sensitivity for detection of human adenovirus (HAdV), compared to an earlier version (RP v1.6). RP v1.7 was designed for detection of species B, C, and E but may show variable detection of species A, D, and F. We sought to evaluate the clinical and analytical performance of RP v1.7 for detection of HAdV in a large pediatric cohort. Respiratory specimens obtained from a tertiary care children's hospital between February 2014 and February 2015 were tested for HAdV by RP v1.7. If the RP v1.7 results were negative for HAdV, then the specimens were reflexed to a HAdV-specific laboratory-developed PCR (LD-PCR) assay for confirmation. A subset of specimens underwent secondary confirmatory testing using another commercially available HAdV PCR assay and a molecular typing assay for species identification. Among 4,750 specimens, a total of 146 specimens (3.1%) were HAdV positive by RP v1.7. HAdV was detected by LD-PCR in an additional 220 specimens that were negative by RP v1.7. Overall, a nearly 5% increase in HAdV detection was observed when RP v1.7-negative specimens were reflexed to LD-PCR testing. RP v1.7 did not detect HAdV with either low viral burden (threshold cycle values of >30) or nonrespiratory species (species A, D, and F), as shown in both clinical and analytic data. While the level of sensitivity of RP v1.7 may be adequate for testing among otherwise healthy children, the decreased sensitivity may be problematic for immunocompromised patients, in whom low levels of HAdV in the respiratory tract may precede systemic infection and require early intervention.
A Method for Amplicon Deep Sequencing of Drug Resistance Genes in Plasmodium falciparum Clinical Isolates from India
Authors: Rao Pavitra N. , Uplekar Swapna , Kayal Sriti , Mallick Prashant K. , Bandyopadhyay Nabamita , Kale Sonal , Singh Om P. , Mohanty Akshaya , Mohanty Sanjib , Wassmer Samuel C. , Carlton Jane M.
Published: March 2016

A major challenge to global malaria control and elimination is early detection and containment of emerging drug resistance. Next-generation sequencing (NGS) methods provide the resolution, scalability, and sensitivity required for high-throughput surveillance of molecular markers of drug resistance. We have developed an amplicon sequencing method on the Ion Torrent PGM platform for targeted resequencing of a panel of six
Plasmodium falciparum
genes implicated in resistance to first-line antimalarial therapy, including artemisinin combination therapy, chloroquine, and sulfadoxine-pyrimethamine. The protocol was optimized using 12 geographically diverse
P. falciparum
reference strains and successfully applied to multiplexed sequencing of 16 clinical isolates from India. The sequencing results from the reference strains showed 100% concordance with previously reported drug resistance-associated mutations. Single-nucleotide polymorphisms (SNPs) in clinical isolates revealed a number of known resistance-associated mutations and other nonsynonymous mutations that have not been implicated in drug resistance. SNP positions containing multiple allelic variants were used to identify three clinical samples containing mixed genotypes indicative of multiclonal infections. The amplicon sequencing protocol has been designed for the benchtop Ion Torrent PGM platform and can be operated with minimal bioinformatics infrastructure, making it ideal for use in countries that are endemic for the disease to facilitate routine large-scale surveillance of the emergence of drug resistance and to ensure continued success of the malaria treatment policy.
Role of Antifungal Susceptibility Testing in Non-Aspergillus Invasive Mold Infections
Authors: Lamoth Frédéric , Damonti Lauro , Alexander Barbara D.
Published: March 2016

No clinical breakpoints are available to delineate antifungal drug efficacy in non-
Aspergillus
invasive mold infections (NAIMIs). In this analysis of 39 NAIMI episodes, the MIC of the first-line antifungal drug was the most important predictor of therapeutic response. For amphotericin B, an MIC of ≤0.5 μg/ml was significantly associated with better 6-week outcomes.
Optimal Cutoff and Accuracy of an IgM Enzyme-Linked Immunosorbent Assay for Diagnosis of Acute Scrub Typhus in Northern Thailand: an Alternative Reference Method to the IgM Immunofluorescence Assay
Authors: Blacksell Stuart D. , Lim Cherry , Tanganuchitcharnchai Ampai , Jintaworn Suthatip , Kantipong Pacharee , Richards Allen L. , Paris Daniel H. , Limmathurotsakul Direk , Day Nicholas P. J.
Published: March 2016

The enzyme-linked immunosorbent assay (ELISA) has been proposed as an alternative serologic diagnostic test to the indirect immunofluorescence assay (IFA) for scrub typhus. Here, we systematically determine the optimal sample dilution and cutoff optical density (OD) and estimate the accuracy of IgM ELISA using Bayesian latent class models (LCMs). Data from 135 patients with undifferentiated fever were reevaluated using Bayesian LCMs. Every patient was evaluated for the presence of an eschar and tested with a blood culture for
Orientia tsutsugamushi
, three different PCR assays, and an IgM IFA. The IgM ELISA was performed for every sample at sample dilutions from 1:100 to 1:102,400 using crude whole-cell antigens of the Karp, Kato, and Gilliam strains of
O. tsutsugamushi
developed by the Naval Medical Research Center. We used Bayesian LCMs to generate unbiased receiver operating characteristic curves and found that the sample dilution of 1:400 was optimal for the IgM ELISA. With the optimal cutoff OD of 1.474 at a sample dilution of 1:400, the IgM ELISA had a sensitivity of 85.7% (95% credible interval [CrI], 77.4% to 86.7%) and a specificity of 98.1% (95% CrI, 97.2% to 100%) using paired samples. For the ELISA, the OD could be determined objectively and quickly, in contrast to the reading of IFA slides, which was both subjective and labor-intensive. The IgM ELISA for scrub typhus has high diagnostic accuracy and is less subjective than the IgM IFA. We suggest that the IgM ELISA may be used as an alternative reference test to the IgM IFA for the serological diagnosis of scrub typhus.
Detection of Pneumocystis jirovecii by Quantitative PCR To Differentiate Colonization and Pneumonia in Immunocompromised HIV-Positive and HIV-Negative Patients
Authors: Fauchier T. , Hasseine L. , Gari-Toussaint M. , Casanova V. , Marty P. M. , Pomares C.
Published: March 2016

Pneumocystis
jirovecii
pneumonia (PCP) is an acute and life-threatening lung disease caused by the fungus
Pneumocystis jirovecii
. The presentation of PCP in HIV-positive patients is well-known and consists of a triad of dyspnea, fever, and cough, whereas the presentation of PCP in HIV-negative patients is atypical and consists of a sudden outbreak, O
2
desaturation, and a rapid lethal outcome without therapy. Despite the availability of direct and indirect identification methods, the diagnosis of PCP remains difficult. The cycle threshold (

C
T

) values obtained by quantitative PCR (qPCR) allow estimation of the fungal burden. The more elevated that the fungal burden is, the higher the probability that the diagnosis is pneumonia. The purposes of the present study were to evaluate the

C
T

values to differentiate colonization and pneumonia in a population of immunocompromised patients overall and patients stratified on the basis of their HIV infection status. Testing of bronchoalveolar lavage (BAL) fluid samples from the whole population of qPCR-positive patients showed a mean

C
T

value for patients with PCP of 28 (95% confidence interval [CI], 26 to 30) and a mean

C
T

value for colonized patients of 35 (95% CI, 34 to 36) (
P
< 10
−3
). For the subgroup of HIV-positive patients, we demonstrated that a

C
T

value below 27 excluded colonization and a

C
T

value above 30 excluded PCP with a specificity of 100% and a sensitivity of 80%, respectively. In the subgroup of HIV-negative patients, we demonstrated that a

C
T

value below 31 excluded colonization and a

C
T

value above 35 excluded PCP with a specificity of 80% and a sensitivity of 80%, respectively. Thus, qPCR of BAL fluid samples is an important tool for the differentiation of colonization and pneumonia in
P. jirovecii
-infected immunocompromised patients and patients stratified on the basis of HIV infection status with different

C
T

values.
A Proline-Rich N-Terminal Region of the Dengue Virus NS3 Is Crucial for Infectious Particle Production
Authors: Gebhard Leopoldo G. , Iglesias Néstor G. , Byk Laura A. , Filomatori Claudia V. , De Maio Federico A. , Gamarnik Andrea V.
Published: March 2016

Dengue virus is currently the most important insect-borne viral human pathogen. Viral nonstructural protein 3 (NS3) is a key component of the viral replication machinery that performs multiple functions during viral replication and participates in antiviral evasion. Using dengue virus infectious clones and reporter systems to dissect each step of the viral life cycle, we examined the requirements of different domains of NS3 on viral particle assembly. A thorough site-directed mutagenesis study based on solvent-accessible surface areas of NS3 revealed that, in addition to being essential for RNA replication, different domains of dengue virus NS3 are critically required for production of infectious viral particles. Unexpectedly, point mutations in the protease, interdomain linker, or helicase domain were sufficient to abolish infectious particle formation without affecting translation, polyprotein processing, or RNA replication. In particular, we identified a novel proline-rich N-terminal unstructured region of NS3 that contains several amino acid residues involved in infectious particle formation. We also showed a new role for the interdomain linker of NS3 in virion assembly. In conclusion, we present a comprehensive genetic map of novel NS3 determinants for viral particle assembly. Importantly, our results provide evidence of a central role of NS3 in the coordination of both dengue virus RNA replication and particle formation.

IMPORTANCE
Dengue virus is an important human pathogen, and its prominence is expanding globally; however, basic aspects of its biology are still unclear, hindering the development of effective therapeutic and prophylactic treatments. Little is known about the initial steps of dengue and other flavivirus particle assembly. This process involves a complex interplay between viral and cellular components, making it an attractive antiviral target. Unpredictably, we identified spatially separated regions of the large NS3 viral protein as determinants for dengue virus particle assembly. NS3 is a multifunctional enzyme that participates in different steps of the viral life cycle. Using reporter systems to dissect different viral processes, we identified a novel N-terminal unstructured region of the NS3 protein as crucial for production of viral particles. Based on our findings, we propose new ideas that include NS3 as a possible scaffold for the viral assembly process.
Both Epistasis and Diversifying Selection Drive the Structural Evolution of the Ebola Virus Glycoprotein Mucin-Like Domain
Authors: Ibeh Neke , Nshogozabahizi Jean Claude , Aris-Brosou Stéphane
Published: March 2016

Throughout the last 3 decades, Ebola virus (EBOV) outbreaks have been confined to isolated areas within Central Africa; however, the 2014 variant reached unprecedented transmission and mortality rates. While the outbreak was still under way, it was reported that the variant leading up to this outbreak evolved faster than previous EBOV variants, but evidence for diversifying selection was undetermined. Here, we test this selection hypothesis and show that while previous EBOV outbreaks were preceded by bursts of diversification, evidence for site-specific diversifying selection during the emergence of the 2014 EBOV clade is weak. However, we show strong evidence supporting an interplay between selection and correlated evolution (epistasis), particularly in the mucin-like domain (MLD) of the EBOV glycoprotein. By reconstructing ancestral structures of the MLD, we further propose a structural mechanism explaining how the substitutions that accumulated between 1918 and 1969 distorted the MLD, while more recent epistatic substitutions restored part of the structure, with the most recent substitution being adaptive. We suggest that it is this complex interplay between weak selection, epistasis, and structural constraints that has shaped the evolution of the 2014 EBOV variant.

IMPORTANCE
The role that selection plays in the emergence of viral epidemics remains debated, particularly in the context of the 2014 EBOV outbreak. Most critically, should such evidence exist, it is generally unclear how this relates to function and increased virulence. Here, we show that the viral lineage leading up to the 2014 outbreak underwent a complex interplay between selection and correlated evolution (epistasis) in a protein region that is critical for immune evasion. We then reconstructed the three-dimensional structure of this domain and showed that the initial mutations in this lineage deformed the structure, while subsequent mutations restored part of the structure. Along this mutational path, the first and last mutations were adaptive, while the intervening ones were epistatic. Altogether, we provide a mechanistic model that explains how selection and epistasis acted on the structural constraints that materialized during the 2014 EBOV outbreak.
Bluetongue Virus NS4 Protein Is an Interferon Antagonist and a Determinant of Virus Virulence
Authors: Ratinier Maxime , Shaw Andrew E. , Barry Gerald , Gu Quan , Di Gialleonardo Luigina , Janowicz Anna , Varela Mariana , Randall Richard E. , Caporale Marco , Palmarini Massimo
Published: March 2016

Bluetongue virus (BTV) is the causative agent of bluetongue, a major infectious disease of ruminants with serious consequences to both animal health and the economy. The clinical outcome of BTV infection is highly variable and dependent on a variety of factors related to both the virus and the host. In this study, we show that the BTV nonstructural protein NS4 favors viral replication in sheep, the animal species most affected by bluetongue. In addition, NS4 confers a replication advantage on the virus in interferon (IFN)-competent primary sheep endothelial cells and immortalized cell lines. We determined that in cells infected with an NS4 deletion mutant (BTV8ΔNS4), there is increased synthesis of type I IFN compared to cells infected with wild-type BTV-8. In addition, using RNA sequencing (RNA-seq), we show that NS4 modulates the host IFN response and downregulates mRNA levels of type I IFN and interferon-stimulated genes. Moreover, using reporter assays and protein synthesis assays, we show that NS4 downregulates the activities of a variety of promoters, such as the cytomegalovirus immediate-early promoter, the IFN-β promoter, and a promoter containing interferon-stimulated response elements (ISRE). We also show that the NS4 inhibitory activity on gene expression is related to its nucleolar localization. Furthermore, NS4 does not affect mRNA splicing or cellular translation. The data obtained in this study strongly suggest that BTV NS4 is an IFN antagonist and a key determinant of viral virulence.

IMPORTANCE
Bluetongue is one of the main infectious diseases of ruminants and is caused by bluetongue virus (BTV), an arthropod-borne virus transmitted from infected to susceptible animals by
Culicoides
biting midges. Bluetongue has a variable clinical outcome that can be related to both virus and host factors. It is therefore critical to understand the interplay between BTV and the host immune responses. In this study, we show that a nonstructural protein of BTV (NS4) is critical to counteract the innate immune response of the host. Infection of cells with a BTV mutant lacking NS4 results in increased synthesis of IFN-β and upregulation of interferon-stimulated genes. In addition, we show that NS4 is a virulence factor for BTV by favoring viral replication in sheep, the animal species most susceptible to bluetongue.
Porcine Reproductive and Respiratory Syndrome Virus Nucleocapsid Protein Interacts with Nsp9 and Cellular DHX9 To Regulate Viral RNA Synthesis
Authors: Liu Long , Tian Jiao , Nan Hao , Tian Mengmeng , Li Yuan , Xu Xiaodong , Huang Baicheng , Zhou Enmin , Hiscox Julian A. , Chen Hongying
Published: March 2016

Porcine reproductive and respiratory syndrome virus (PRRSV) nucleocapsid (N) protein is the main component of the viral capsid to encapsulate viral RNA, and it is also a multifunctional protein involved in the regulation of host cell processes. Nonstructural protein 9 (Nsp9) is the RNA-dependent RNA polymerase that plays a critical role in viral RNA transcription and replication. In this study, we demonstrate that PRRSV N protein is bound to Nsp9 by protein-protein interaction and that the contacting surface on Nsp9 is located in the two predicted α-helixes formed by 48 residues at the C-terminal end of the protein. Mutagenesis analyses identified E646, E608, and E611 on Nsp9 and Q85 on the N protein as the pivotal residues participating in the N-Nsp9 interaction. By overexpressing the N protein binding fragment of Nsp9 in infected Marc-145 cells, the synthesis of viral RNAs, as well as the production of infectious progeny viruses, was dramatically inhibited, suggesting that Nsp9-N protein association is involved in the process of viral RNA production. In addition, we show that PRRSV N interacts with cellular RNA helicase DHX9 and redistributes the protein into the cytoplasm. Knockdown of DHX9 increased the ratio of short subgenomic mRNAs (sgmRNAs); in contrast, DHX9 overexpression benefited the synthesis of longer sgmRNAs and the viral genomic RNA (gRNA). These results imply that DHX9 is recruited by the N protein in PRRSV infection to regulate viral RNA synthesis. We postulate that N and DHX9 may act as antiattenuation factors for the continuous elongation of nascent transcript during negative-strand RNA synthesis.

IMPORTANCE
It is unclear whether the N protein of PRRSV is involved in regulation of the viral RNA production process. In this report, we demonstrate that the N protein of the arterivirus PRRSV participates in viral RNA replication and transcription through interacting with Nsp9 and its RdRp and recruiting cellular RNA helicase to promote the production of longer viral sgmRNAs and gRNA. Our data here provide some new insights into the discontinuous to continuous extension of PRRSV RNA synthesis and also offer a new potential anti-PRRSV strategy targeting the N-Nsp9 and/or N-DHX9 interaction.
Mutagenesis of Coronavirus nsp14 Reveals Its Potential Role in Modulation of the Innate Immune Response
Authors: Becares Martina , Pascual-Iglesias Alejandro , Nogales Aitor , Sola Isabel , Enjuanes Luis , Zuñiga Sonia
Published: March 2016

Coronavirus (CoV) nonstructural protein 14 (nsp14) is a 60-kDa protein encoded by the replicase gene that is part of the replication-transcription complex. It is a bifunctional enzyme bearing 3′-to-5′ exoribonuclease (ExoN) and guanine-N7-methyltransferase (N7-MTase) activities. ExoN hydrolyzes single-stranded RNAs and double-stranded RNAs (dsRNAs) and is part of a proofreading system responsible for the high fidelity of CoV replication. nsp14 N7-MTase activity is required for viral mRNA cap synthesis and prevents the recognition of viral mRNAs as “non-self” by the host cell. In this work, a set of point mutants affecting different motifs within the ExoN domain of nsp14 was generated, using transmissible gastroenteritis virus as a model of
Alphacoronavirus
. Mutants lacking ExoN activity were nonviable despite being competent in both viral RNA and protein synthesis. A specific mutation within zinc finger 1 (ZF-C) led to production of a viable virus with growth and viral RNA synthesis kinetics similar to that of the parental virus. Mutant recombinant transmissible gastroenteritis virus (TGEV) ZF-C (rTGEV-ZF-C) caused decreased cytopathic effect and apoptosis compared with the wild-type virus and reduced levels of dsRNA accumulation at late times postinfection. Consequently, the mutant triggered a reduced antiviral response, which was confirmed by evaluating different stages of the dsRNA-induced antiviral pathway. The expression of beta interferon (IFN-β), tumor necrosis factor (TNF), and interferon-stimulated genes in cells infected with mutant rTGEV-ZF-C was reduced compared to the levels seen with the parental virus. Overall, our data revealed a potential role for CoV nsp14 in modulation of the innate immune response.

IMPORTANCE
The innate immune response is the first line of antiviral defense that culminates in the synthesis of interferon and proinflammatory cytokines to control viral replication. CoVs have evolved several mechanisms to counteract the innate immune response at different levels, but the role of CoV-encoded ribonucleases in preventing activation of the dsRNA-induced antiviral response has not been described to date. The introduction of a mutation in zinc finger 1 of the ExoN domain of nsp14 led to production of a virus that induced a weak antiviral response, most likely due to the accumulation of lower levels of dsRNA in the late phases of infection. These observations allowed us to propose a novel role for CoV nsp14 ExoN activity in counteracting the antiviral response, which could serve as a novel target for the design of antiviral strategies.
Comparative Cell Biology and Evolution of Annexins in Diplomonads
Authors: Einarsson Elin , Ástvaldsson Ásgeir , Hultenby Kjell , Andersson Jan O. , Svärd Staffan G. , Jerlström-Hultqvist Jon
Published: March 2016

Annexins are proteins that associate with phospholipids in a Ca
2+
-dependent fashion. These proteins have been intensely studied in animals and plants because of their importance in diverse cellular processes, yet very little is known about annexins in single-celled eukaryotes, which represent the largest diversity of organisms. The human intestinal parasite
Giardia intestinalis
is known to have more annexins than humans, and they contribute to its pathogenic potential. In this study, we investigated the annexin complement in the salmon pathogen
Spironucleus salmonicida
, a relative of
G. intestinalis
. We found that
S. salmonicida
has a large repertoire of annexins and that the gene family has expanded separately across diplomonads, with members showing sequence diversity similar to that seen across kingdom-level groups such as plants and animals.
S. salmonicida
annexins are prominent components of the cytoskeleton and membrane. Two annexins are associated with a previously unrecognized structure in the anterior of the cell.
Estimated incidence and risk factors of sudden unexpected death
Authors: Lewis Mary Elizabeth , Lin Feng-Chang , Nanavati Parin , Mehta Neil , Mounsey Louisa , Nwosu Anthony , Pursell Irion , Chung Eugene H , Mounsey J Paul , Simpson Ross J
Published: March 2016

Objective

In this manuscript, we estimate the incidence and identify risk factors for sudden unexpected death in a socioeconomically and racially diverse population in one county in North Carolina. Estimates of the incidence and risk factors contributing to sudden death vary widely. The Sudden Unexpected Death in North Carolina (SUDDEN) project is a population-based investigation of the incidence and potential causes of sudden death.

Methods

From 3 March 2013 to 2 March 2014, all out-of-hospital deaths in Wake County, North Carolina, were screened to identify presumed sudden unexpected death among free-living residents between the ages of 18 and 64 years. Death certificate, public and medical records were reviewed and adjudicated to confirm sudden unexpected death cases.

Results

Following adjudication, 190 sudden unexpected deaths including 122 men and 68 women were identified. Estimated incidence was 32.1 per 100 000 person-years overall: 42.7 among men and 22.4 among women. The majority of victims were white, unmarried men over age 55 years, with unwitnessed deaths at home. Hypertension and dyslipidaemia were common in men and women. African-American women dying from sudden unexpected death were over-represented. Women who were under age 55 years with coronary disease accounted for over half of female participants with coronary artery disease.

Conclusions

The overall estimated incidence of sudden unexpected death may account for approximately 10% of all deaths classified as ‘natural’. Women have a lower estimated incidence of sudden unexpected death than men. However, we found no major differences in age or comorbidities between men and women. African-Americans and young women with coronary disease are at risk for sudden unexpected death.
Ten‐minute chat
Authors: Peel Barbara
Published: March 2016

Barbara Peel is an animal nurse at Goddard Veterinary Group's Collier Row practice in Romford. Her 35‐year career began rather unexpectedly and was the result of a heavy fall of snow.
Feed intake, growth, and behavioral assessment of mink fed a clam-based diet
Authors: Campbell Dana L.M. , Link Jane E. , Lester-Saenz Amber H. , Bursian Steven J.
Published: March 2016

The objective of this study was to assess the acceptance and safety of two species of food-grade clams, ocean quahog (Arctica islandica) and Atlantic surf clam (Spisula solidissima), for consumption by fur-farmed American mink (Neovison vison). These clams contained thiaminase that can lead to weight loss, paralysis, and death. Across 19 wk, 39 black, adult female mink divided into three groups were fed a standard chicken-based control diet, 10% clam diet, or 20% clam diet and assessed for feed consumption rates, body weights, locomotor stereotypic behavior, overall activity levels, and ability to complete a novel tunnel climbing task. The 20% clam group consumed significantly less metabolizable energy compared with the control group, but there was no significant difference in weight gain between groups. There were some inconsistent group differences in stereotypy and overall activity with all groups showing significant behavioral differences across study weeks; possibly related to ambient weather and photoperiod. There were no significant group differences in their ability to complete the tunnel task. Based on our results, no negative effect of clam consumption was observed in adult female mink, but further studies are needed to determine clam safety to both male and female mink from all life stages.
Blood haemogram in Ovis aries and Capra hyrcus: effect of storage time
Authors: Fazio Francesco , Giangrosso Giuseppe , Marafioti Simona , Zanghì Elisa , Arfuso Francesca , Piccione Giuseppe
Published: March 2016

The aim of the present study was to assess the effect of storage time at +4 °C on haematological profile in goat (n = 25) and sheep (n = 25). After collection, blood samples were immediately analyzed and then divided into four aliquots that were stored at 4 °C and tested at 24 h (T1), 48 h (T2), 72 h (T3), and 1 wk (T4), respectively. One-way repeated-measures analysis of variance (ANOVA) was used to determine statistically significant effect of storage conditions both in goats and in sheep. Our results showed that among the two species studied, goats showed highest blood stability after refrigeration at +4 °C. In goats, all hematological parameters, except PLT, showed no significant changes during all days of monitoring with respect to basal values (T0). In sheep, no significantly effect of storage time on RBC and WBC levels were found, whereas the other hematological parameters change significantly over the time. Our findings suggest that the blood storage time reported for goat may not be applied to sheep’s blood, which underscore the differences between these two species that are erroneously considered similar.
Caveolin-1 is a checkpoint regulator in hypoxia-induced astrocyte apoptosis via Ras/Raf/ERK pathway
Authors: Xu Lili , Wang Liumin , Wen Zhuoyu , Wu Li , Jiang Yongjun , Yang Lian , Xiao Lulu , Xie Yi , Ma Minmin , Zhu Wusheng , Ye Ruidong , Liu Xinfeng
Published: March 2016

Astrocytes, the most numerous cells in the human brain, play a central role in the metabolic homeostasis following hypoxic injury. Caveolin-1 (Cav-1), a transmembrane scaffolding protein, has been shown to converge prosurvival signaling in the central nerve system. The present study aimed to investigate the role of Cav-1 in the hypoxia-induced astrocyte injury. We also examined how Cav-1 alleviates apoptotic astrocyte death. To this end, primary astrocytes were exposed to oxygen-glucose deprivation (OGD) for 6 h and a subsequent 24-h reoxygenation to mimic hypoxic injury. OGD significantly reduced Cav-1 expression. Downregulation of Cav-1 using Cav-1 small interfering RNA dramatically worsened astrocyte cell damage and impaired cellular glutamate uptake after OGD, whereas overexpression of Cav-1 with Cav-1 scaffolding domain peptide attenuated OGD-induced cell apoptosis. Mechanistically, the expressions of Ras-GTP, phospho-Raf, and phospho-ERK were sequestered in Cav-1 small interfering RNA-treated astrocytes, yet were stimulated after supplementation with caveolin peptide. MEK/ERK inhibitor U0126 remarkably blocked the Cav-1-induced counteraction against apoptosis following hypoxia, indicating Ras/Raf/ERK pathway is required for the Cav-1's prosurvival role. Together, these findings support Cav-1 as a checkpoint for the in hypoxia-induced astrocyte apoptosis and warrant further studies targeting Cav-1 to treat hypoxic-ischemic brain injury.
Macrophage-derived IL-6 contributes to ANG II-mediated angiotensinogen stimulation in renal proximal tubular cells
Authors: O'Leary Ryan , Penrose Harrison , Miyata Kayoko , Satou Ryousuke
Published: March 2016

The development of ANG II-dependent hypertension involves increased infiltration of macrophages (MΦ) and T cells into the kidney and the consequent elevation of intrarenal cytokines including IL-6, which facilitates the progression of hypertension and associated kidney injury. Intrarenal renin-angiotensin system (RAS) activation, including proximal tubular angiotensinogen (AGT) stimulation, has also been regarded as a cardinal mechanism contributing to these diseases. However, the interaction between immune cells and intrarenal RAS activation has not been fully delineated. Therefore, the present study investigated whether ANG II-treated MΦ induce AGT upregulation in renal proximal tubular cells (PTCs). MΦ were treated with 0–10−6M ANG II for up to 48 h. PTCs were incubated with the collected medium from MΦ. In ANG II-treated MΦ, IL-6 mRNA and protein levels were increased (1.86 ± 0.14, protein level, ratio to control); moreover, IL-6 levels were higher than TNF-α and IL-1β in culture medium isolated from ANG II-treated MΦ. Elevated AGT expression (1.69 ± 0.04, ratio to control) accompanied by phosphorylated STAT3 were observed in PTCs that received culture medium from ANG II-treated MΦ. The addition of a neutralizing IL-6 antibody to the collected medium attenuated phosphorylation of STAT3 and AGT augmentation in PTCs. Furthermore, a JAK2 inhibitor also suppressed STAT3 phosphorylation and AGT augmentation in PTCs. These results demonstrate that ANG II-induced IL-6 elevation in MΦ enhances activation of the JAK-STAT pathway and consequent AGT upregulation in PTCs, suggesting involvement of an immune response in driving intrarenal RAS activity.
LRP predicts smooth pursuit eye movement onset during the ocular tracking of self-generated movements
Authors: Chen Jing , Valsecchi Matteo , Gegenfurtner Karl R.
Published: March 2016

Several studies have indicated that human observers are very efficient at tracking self-generated hand movements with their gaze, yet it is not clear whether this is simply a by-product of the predictability of self-generated actions or if it results from a deeper coupling of the somatomotor and oculomotor systems. In a first behavioral experiment we compared pursuit performance as observers either followed their own finger or tracked a dot whose motion was externally generated but mimicked their finger motion. We found that even when the dot motion was completely predictable in terms of both onset time and kinematics, pursuit was not identical to that produced as the observers tracked their finger, as evidenced by increased rate of catch-up saccades and by the fact that in the initial phase of the movement gaze was lagging behind the dot, whereas it was ahead of the finger. In a second experiment we recorded EEG in the attempt to find a direct link between the finger motor preparation, indexed by the lateralized readiness potential (LRP) and the latency of smooth pursuit. After taking into account finger movement onset variability, we observed larger LRP amplitudes associated with earlier smooth pursuit onset across trials. The same held across subjects, where average LRP onset correlated with average eye latency. The evidence from both experiments concurs to indicate that a strong coupling exists between the motor systems leading to eye and finger movements and that simple top-down predictive signals are unlikely to support optimal coordination.
Biophysical constraints on lateral inhibition in the olfactory bulb
Authors: McIntyre Alexa B. R. , Cleland Thomas A.
Published: March 2016

The mitral cells (MCs) of the mammalian olfactory bulb (OB) constitute one of two populations of principal neurons (along with middle/deep tufted cells) that integrate afferent olfactory information with top-down inputs and intrinsic learning and deliver output to downstream olfactory areas. MC activity is regulated in part by inhibition from granule cells, which form reciprocal synapses with MCs along the extents of their lateral dendrites. However, with MC lateral dendrites reaching over 1.5 mm in length in rats, the roles of distal inhibitory synapses pose a quandary. Here, we systematically vary the properties of a MC model to assess the capacity of inhibitory synaptic inputs on lateral dendrites to influence afferent information flow through MCs. Simulations using passivized models with varying dendritic morphologies and synaptic properties demonstrated that, even with unrealistically favorable parameters, passive propagation fails to convey effective inhibitory signals to the soma from distal sources. Additional simulations using an active model exhibiting action potentials, subthreshold oscillations, and a dendritic morphology closely matched to experimental values further confirmed that distal synaptic inputs along the lateral dendrite could not exert physiologically relevant effects on MC spike timing at the soma. Larger synaptic conductances representative of multiple simultaneous inputs were not sufficient to compensate for the decline in signal with distance. Reciprocal synapses on distal MC lateral dendrites may instead serve to maintain a common fast oscillatory clock across the OB by delaying spike propagation within the lateral dendrites themselves.
Meckel's Diverticulum in Crohn's Disease
Authors: Freeman Hugh J
Published: March 2016

Meckel's diverticulum is a congenital abnormality of the distal ileum associated with failed vitelline duct closure. Detailed pathological studies have estimated its frequency to be about 2% of the general population, and it has been anecdotally recorded in patients with Crohn's disease. Most patients with Crohn's disease have imaging studies of the small intestine during the course of their disease, and often, an intestinal resection. Thus, it seems possible to estimate the prevalence of Meckel's diverticula in Crohn's disease. In addition, patient characteristics may be important, especially if management of Crohn's disease is altered. Of 877 patients with Crohn's disease, 10 (about 1%) had a Meckel's diverticulum diagnosed, including six men and four women. All were diagnosed with Crohn's disease before age 50 years and seven were diagnosed before age 30 years. There were five with ileocolonic disease, two with colon-only disease and three with ileum-only disease. The clinical behaviour of five patients could be classified as penetrating and two as stricturing. A total of 311 patients had an ileocolonic resection, including eight (about 2%) with a Meckel's diverticulum. In contrast to some case reports, no heterotopic mucosa was detected and the Meckel's diverticulum was incidental and, apparently, an unexpected finding. In each case, the diverticulum was not involved with Crohn's disease but was included in the ileal resection. These results suggest that the overall prevalence of a Meckel's diverticulum is not increased in Crohn's disease but may result in resection of additional small intestine.
In vivo gluten challenge in coeliac disease
Authors: Ellis HJ , Ciclitira PJ
Published: March 2016

In vivo gluten challenge has been used since the early 1950s to study the role of cereal fractions in celiac disease. While early studies relied on crude indicators of celiac toxicity, the advent of jejunal biopsy and sophisticated immunohistochemical techniques has allowed accurate studies to be performed. Studies to determine the nature of the cereal component that is toxic to patients with celiac disease have concentrated on wheat because of its nutritional importance. A number of in vitro studies indicated the presence of one or more celiac-activating epitopes with theN-terminus of the A-gliadin molecule. In vivo challenge with three synthetic peptides subsequently indicated the toxicity of a peptide corresponding to amino acids 31 to 49 of A-gliadin. In vivo gluten challenge is the gold standard for the assessment of celiac toxicity; however, jejunal biopsy is a relatively invasive procedure, thus, other methods have been investigated. Direct infusion of the rectum with gluten has been shown to result in an increase in mucosal intraepithelial lymphocytes, occurring only in celiac patients. This method has been used to study the celiac toxicity of gliadin subfractions. The in vitro technique of small intestinal biopsy organ culture is also a useful tool and appears to give the same results as in vivo challenge. The importance of tiny amounts of gliadin in the diet, such as that which occurs in wheat starch, has been studied by in vivo challenge; this technique has clarified the position of oats in the gluten-free diet. Several studies suggest that this cereal may be included in the diet of most adult celiac patients. Studies of the transport of gliadin across the enterocyte following ingestion or challenge suggest that gliadin may be metabolized by a different pathway in celiac disease. This could result in an abnormal presentation to the immune system, triggering a pathogenic rather than a tolerogenic response.
Gastrointestinal Mucormycosis in a Renal Transplant Patient
Authors: Tinmouth Jill , Baker Jeffrey , Gardiner Geoffrey
Published: March 2016

The clinical course and management of a rare case of gastrointestinal mucormycosis occurring in a renal transplant patient are presented. The diagnosis was made on pathological examination of surgically resected tissue from the colon, spleen and stomach. The patient did not survive the infection. To the best of our knowledge, this is the 11th reported case of gastrointestinal mucormycosis in a solid organ transplant patient. The pathophysiology, incidence and prognosis of this disease are discussed.
Topographic Mapping of Collagenous Gastritis
Authors: Freeman Hugh J
Published: March 2016

A 74-year-old woman was investigated for abdominal pain and diarrhea. Endoscopic examinations including biopsies of the stomach and colon demonstrated the typical subepithelial deposits characteristic of collagenous gastritis and collagenous colitis. Histochemical and ultrastructural methods confirmed the presence of collagen in the subepithelial deposits. The topographic distribution of these collagen deposits and their relationship to the inflammatory process in the stomach were then defined by endoscopic mapping and multiple site biopsies of the mucosa in the gastric body and antrum. These studies indicate that collagenous gastritis not only is distinctive, but also is a far more extensive and diffuse inflammatory process than has previously been appreciated.
Heparin‐Induced Hepatotoxicity
Authors: AL-Mekhaizeem Khalid A , Sherker Averell H
Published: March 2016

Heparin‐induced hepatotoxicity is well described in the literature, but rarely recognized clinically. Two cases were recently encountered. In the first case, elevated aminotransferase levels occurred after four days of heparin therapy. In the second case, enzyme levels increased after only 8 h of heparin treatment. To the authors′ knowledge, this short time interval between the administration of heparin and liver enzyme elevations has not been described. The objective of this report is to increase the clinical awareness of this interesting and under‐recognized biochemical observation.
Screening Strategies for Colorectal Cancer: A Systematic Review of the Evidence
Authors: McLeod Robin S ,   members of the Canadian Task Force on Preventive Health Care
Published: March 2016

BACKGROUND: Colorectal cancer is the third most common cancer in Canada. It is well recognized that there are improved survival rates if the disease is treated in its early stages, and indeed this may be a preventable disease. This paper systematically reviews the effectiveness of specific screening techniques for colorectal cancer in asymptomatic individuals at normal or above average risk.

METHODS: MEDLINE was searched for articles published between January 1966 and January 2001 by using the MESH terms 'screening' and 'colorectal neoplasia'. The reference sections of review articles published before January 2001 were checked, and content experts were surveyed. The evidence was evaluated using the standardized methodology of the Canadian Task Force on Preventive Health Care.

RSULTS AND DISCUSSION: For individuals at normal risk, there is evidence to support the use of annual or biennial fecal occult blood testing and flexible sigmoidoscopy for asymptomatic individuals over age 50 years. The evidence regarding whether only one or both of fecal occult blood testing and sigmoidoscopy should be performed is unclear, as is the evidence regarding the use of colonoscopy as an initial screen. For individuals at above average risk, the evidence supports either genetic testing or flexible sigmoidoscopy of individuals at risk in familial adenomatous polyposis kindreds, and screening with colonoscopy of patients in kindreds with hereditary nonpolyposis colon cancer. The evidence regarding colonoscopy for individuals who have a family history of colorectal polyps or cancer but do not fit the criteria for hereditary nonpolyposis colon cancer is unclear. Development of better risk stratification for screening is a high research priority, and further research, including randomized, controlled trials, into the effectiveness and feasibility of other screening modalities is necessary.