title: Silver Sucrose Octasulfate (IASOS™) as a Valid Active Ingredient into a Novel Vaginal Gel against Human Vaginal Pathogens: In Vitro Antimicrobial Activity Assessment

text: This in vitro study assessed the antimicrobial properties of a novel octasilver salt of Sucrose Octasulfate (IASOS) as well as of an innovative vaginal gel containing IASOS (SilSOS Femme), against bacterial and yeast pathogens isolated from human clinical cases of symptomatic vaginal infections. In BHI and LAPT culture media, different ionic silver concentrations and different pHs were tested. IASOS exerted a strong antimicrobial activity towards all the pathogens tested in both culture media. The results demonstrated that salts and organic compounds present in the culture media influenced IASOS efficacy only to a moderate extent. Whereas comparable MBCs (Minimal Bactericidal Concentrations) were observed for G. vaginalis (10 mg/L Ag(+)), E. coli and E. aerogenes (25 mg/L Ag(+)) in both media, higher MBCs were found for S. aureus and S. agalactiae in LAPT cultures (50 mg/L Ag(+) versus 25 mg/L Ag(+)). No minimal concentration totally inhibiting the growth of C. albicans was found. Nevertheless, in both media at the highest ionic silver concentrations (50–200 mg/L Ag(+)), a significant 34–52% drop in Candida growth was observed. pH differently affected the antimicrobial properties of IASOS against bacteria or yeasts; however, a stronger antimicrobial activity at pH higher than the physiological pH was generally observed. It can be therefore concluded that IASOS exerts a bactericidal action against all the tested bacteria and a clear fungistatic action against C. albicans. The antimicrobial activity of the whole vaginal gel SilSOS Femme further confirmed the antimicrobial activity of IASOS. Overall, our findings support IASOS as a valid active ingredient into a vaginal gel.


title: Uncomplicated Plasmodium vivax malaria in pregnancy associated with mortality from acute respiratory distress syndrome

text: The association between severe malaria and Plasmodium vivax species is contentious. On the Thai-Myanmar border, all pregnant women are followed systematically with active weekly malaria screening. Over a 27-year period of providing antenatal care, 48,983 have been prospectively followed until pregnancy outcome (miscarriage or delivery) and 4,298 women have had P. vivax detected at least once. Reported here is the first known P. vivax-associated death amongst these women. The initial patient presentation was of uncomplicated P. vivax (0.5% parasitaemia) in a term, multigravida woman who responded rapidly to oral artesunate and mefloquine treatment, clearing her blood stage parasites within 48 hours. The patient appeared well, was ambulatory and due to be discharged but became unwell with acute respiratory distress syndrome (ARDS) requiring ventilation three days (67 hours) into treatment. Despite induction and delivery of a stillborn foetus, ventilatory requirements increased and the patient died on day 7. The patient had a low body mass index. Sensitive detection with nested PCR confirmed only the presence of P. vivax species and concomitant infections such as tuberculosis and human immunodeficiency virus (HIV) were also ruled out. The contemporaneous treatment of acute uncomplicated P. vivax and the onset of ARDS on day 3 in this patient implies a possible but unconfirmed association with death in this patient. Assuming this death was caused by P. vivax, the risk of ARDS-related maternal mortality in this setting did not differ significantly between Plasmodium falciparum and P. vivax (0.24 per 1,000 (1/4,158) versus 0.23 per 1,000 (1/4,298), contrary to the increased risk of maternal mortality from P. falciparum compared to P. vivax, 2.89 per 1,000 (12/4,158) versus 0.23 per 1,000 (1/4,298), P = 0.003.


title: Using HIV Networks to Inform Real Time Prevention Interventions

text: OBJECTIVE: To reconstruct the local HIV-1 transmission network from 1996 to 2011 and use network data to evaluate and guide efforts to interrupt transmission. DESIGN: HIV-1 pol sequence data were analyzed to infer the local transmission network. METHODS: We analyzed HIV-1 pol sequence data to infer a partial local transmission network among 478 recently HIV-1 infected persons and 170 of their sexual and social contacts in San Diego, California. A transmission network score (TNS) was developed to estimate the risk of HIV transmission from a newly diagnosed individual to a new partner and target prevention interventions. RESULTS: HIV-1 pol sequences from 339 individuals (52.3%) were highly similar to sequences from at least one other participant (i.e., clustered). A high TNS (top 25%) was significantly correlated with baseline risk behaviors (number of unique sexual partners and insertive unprotected anal intercourse (p = 0.014 and p = 0.0455, respectively) and predicted risk of transmission (p<0.0001). Retrospective analysis of antiretroviral therapy (ART) use, and simulations of ART targeted to individuals with the highest TNS, showed significantly reduced network level HIV transmission (p<0.05). CONCLUSIONS: Sequence data from an HIV-1 screening program focused on recently infected persons and their social and sexual contacts enabled the characterization of a highly connected transmission network. The network-based risk score (TNS) was highly correlated with transmission risk behaviors and outcomes, and can be used identify and target effective prevention interventions, like ART, to those at a greater risk for HIV-1 transmission.


title: Interactome Profile of the Host Cellular Proteins and the Nonstructural Protein 2 of Porcine Reproductive and Respiratory Syndrome Virus

text: The nonstructural protein 2 (NSP2) is considered to be one of crucial viral proteins in the replication and pathogenesis of porcine reproductive and respiratory syndrome virus (PRRSV). In the present study, the host cellular proteins that interact with the NSP2 of PRRSV were immunoprecipitated with anti-Myc antibody from the MARC-145 cells infected by a recombinant PRRSV with 3xMyc tag insertion in its NSP2-coding region, and then 285 cellular proteins interacting with NSP2 were identified by LC-MS/MS. The Gene Ontology and enriched KEGG Pathway bioinformatics analyses indicated that the identified proteins could be assigned to different subcellular locations and functional classes. Functional analysis of the interactome profile highlighted cellular pathways associated with infectious disease, translation, immune system, nervous system and signal transduction. Two interested cellular proteins–BCL2-associated athanogene 6 (BAG6) and apoptosis-inducing factor 1 (AIF1) which may involve in transporting of NSP2 to Endoplasmic reticulum (ER) or PRRSV-driven apoptosis were validated by Western blot. The interactome data between PRRSV NSP2 and cellular proteins contribute to the understanding of the roles of NSP2 in the replication and pathogenesis of PRRSV, and also provide novel cellular target proteins for elucidating the associated molecular mechanisms of the interaction of host cellular proteins with viral proteins in regulating the viral replication.


title: A comparative study of experimental mouse models of central nervous system demyelination

text: Several mouse models of multiple sclerosis (MS) are now available. We have established a mouse model, in which ocular infection with a recombinant HSV-1 that expresses murine IL-2 constitutively (HSV-IL-2) causes CNS demyelination in different strains of mice. This model differs from most other models in that it represents a mixture of viral and immune triggers. In the present study, we directly compared MOG(35–55), MBP(35–47), and PLP(190–209) models of EAE with our HSV-IL-2-induced MS model. Mice with HSV-IL-2-induced and MOG-induced demyelinating diseases demonstrated a similar pattern and distribution of demyelination in their brain, spinal cord, and optic nerves. In contrast, no demyelination was detected in the optic nerves of MBP- and PLP-injected mice. IFN-β injections significantly reduced demyelination in brains of all groups, in the spinal cords of the MOG and MBP groups, and completely blocked it in the spinal cords of the PLP and HSV-IL-2 groups as well as in optic nerves of MOG and HSV-IL-2 groups. In contrast to IFN-β treatment, IL-12p70 protected the HSV-IL-2 group from demyelination, while IL-4 was not effective at all in preventing demyelination. MOG-injected mice showed clinical signs of paralysis and disease-related mortality whereas mice in the other treatment groups did not. Collectively, the results indicate that the HSV-IL-2 model and the MOG model complement each other and, together, provide unique insights into the heterogeneity of human MS.


title: Retrospective diagnosis of a famous historical figure: ontological, epistemic, and ethical considerations

text: The aim of this essay is to elaborate philosophical and ethical underpinnings of posthumous diagnosis of famous historical figures based on literary and artistic products, or commonly called retrospective diagnosis. It discusses ontological and epistemic challenges raised in the humanities and social sciences, and attempts to systematically reply to their criticisms from the viewpoint of clinical medicine, philosophy of medicine, particularly the ontology of disease and the epistemology of diagnosis, and medical ethics. The ontological challenge focuses on the doubt about the persistence of a disease over historical time, whereas the epistemic challenge disputes the inaccessibility of scientific verification of a diagnosis in the past. I argue that the critics are in error in conflating the taxonomy of disease (nosology) and the act of diagnosing a patient. Medical diagnosis is fundamentally a hypothesis-construction and an explanatory device that can be generated under various degrees of uncertainty and limited amount of information. It is not an apodictic judgment (true or false) as the critics presuppose, but a probabilistic (Bayesian) judgment with varying degrees of plausibility under uncertainty. In order to avoid this confusion, I propose that retrospective diagnosis of a historical figure be syndromic without identifying underlying disease, unless there is justifiable reason for such specification. Moreover it should be evaluated not only from the viewpoint of medical science but also in a larger context of the scholarship of the humanities and social sciences by its overall plausibility and consistency. On the other hand, I will endorse their concerns regarding the ethics and professionalism of retrospective diagnosis, and call for the need for situating such a diagnosis in an interdisciplinary scope and the context of the scholarship of the historical figure. I will then enumerate several important caveats for interdisciplinary retrospective diagnosis using an example of the retrospective diagnosis of Socrates for his life-long intermittent neurologic symptoms. Finally, I will situate the present argument in a larger context of the major debate among the historians of medicine and paleopathologists, and discuss the similarities and differences.


title: Core Self-Evaluations Mediate the Associations of Dispositional Optimism and Life Satisfaction

text: BACKGROUND: Positive traits, such as life satisfaction, optimism, and core self-evaluation (CSE), have garnered increasing attention from researchers and professionals. However, the trilateral relationship among them remains unclear. OBJECTIVE: This study examines the effect of dispositional optimism on life satisfaction and primarily verified the mediator role of CSEs. METHODS: Six hundred thirty college students from two general universities completed a questionnaire packet containing life orientation test–revised (LOT–R), core self-evaluations, and satisfaction with life scale. Confirmatory factor analysis (CFA) was conducted to assess the dimension of LOT–R. Bootstrap was used in structural equation modeling to analyze mediation effect. RESULTS: Results revealed that dispositional optimism and core self-evaluations were significantly correlated with life satisfaction. CFA identified the bidimensional structure of dispositional optimism. SEM indicated that core self-evaluations partially mediated the effect of dispositional optimism on life satisfaction. The final model also revealed significant paths from optimism and pessimism to life satisfaction through core-self evaluations. CONCLUSION: The findings extended prior studies and shed light on how dispositional optimism influences life satisfaction. This study provides valuable evidence on how to promote the life satisfaction of human beings in positive psychology. A further study can fully explore the relationship among them in multi-cultural follow-up studies.


title: Management of sepsis in neutropenic patients: 2014 updated guidelines from the Infectious Diseases Working Party of the German Society of Hematology and Medical Oncology (AGIHO)

text: Sepsis is a major cause of mortality during the neutropenic phase after intensive cytotoxic therapies for malignancies. Improved management of sepsis during neutropenia may reduce the mortality of cancer therapies. Clinical guidelines on sepsis treatment have been published by others. However, optimal management may differ between neutropenic and non-neutropenic patients. Our aim is to give evidence-based recommendations for haematologist, oncologists and intensive care physicians on how to manage adult patients with neutropenia and sepsis.


title: Estimated Effects of Projected Climate Change on the Basic Reproductive Number of the Lyme Disease Vector Ixodes scapularis

text: Background: The extent to which climate change may affect human health by increasing risk from vector-borne diseases has been under considerable debate. Objectives: We quantified potential effects of future climate change on the basic reproduction number (R(0)) of the tick vector of Lyme disease, Ixodes scapularis, and explored their importance for Lyme disease risk, and for vector-borne diseases in general. Methods: We applied observed temperature data for North America and projected temperatures using regional climate models to drive an I. scapularis population model to hindcast recent, and project future, effects of climate warming on R(0). Modeled R(0) increases were compared with R(0) ranges for pathogens and parasites associated with variations in key ecological and epidemiological factors (obtained by literature review) to assess their epidemiological importance. Results: R(0) for I. scapularis in North America increased during the years 1971–2010 in spatio-temporal patterns consistent with observations. Increased temperatures due to projected climate change increased R(0) by factors (2–5 times in Canada and 1.5–2 times in the United States), comparable to observed ranges of R(0) for pathogens and parasites due to variations in strains, geographic locations, epidemics, host and vector densities, and control efforts. Conclusions: Climate warming may have co-driven the emergence of Lyme disease in northeastern North America, and in the future may drive substantial disease spread into new geographic regions and increase tick-borne disease risk where climate is currently suitable. Our findings highlight the potential for climate change to have profound effects on vectors and vector-borne diseases, and the need to refocus efforts to understand these effects. Citation: Ogden NH, Radojević M, Wu X, Duvvuri VR, Leighton PA, Wu J. 2014. Estimated effects of projected climate change on the basic reproductive number of the Lyme disease vector Ixodes scapularis. Environ Health Perspect 122:631–638; http://dx.doi.org/10.1289/ehp.1307799


title: NGS Nominated CELA1, HSPG2, and KCNK5 as Candidate Genes for Predisposition to Balkan Endemic Nephropathy

text: Balkan endemic nephropathy (BEN) is a familial chronic tubulointerstitial disease with insidious onset and slow progression leading to terminal renal failure. The results of molecular biological investigations propose that BEN is a multifactorial disease with genetic predisposition to environmental risk agents. Exome sequencing of 22 000 genes with Illumina Nextera Exome Enrichment Kit was performed on 22 DNA samples (11 Bulgarian patients and 11 Serbian patients). Software analysis was performed via NextGene, Provean, and PolyPhen. The frequency of all annotated genetic variants with deleterious/damaging effect was compared with those of European populations. Then we focused on nonannotated variants (with no data available about them and not found in healthy Bulgarian controls). There is no statistically significant difference between annotated variants in BEN patients and European populations. From nonannotated variants with more than 40% frequency in both patients' groups, we nominated 3 genes with possible deleterious/damaging variants—CELA1, HSPG2, and KCNK5. Mutant genes (CELA1, HSPG2, and KCNK5) in BEN patients encode proteins involved in basement membrane/extracellular matrix and vascular tone, tightly connected to process of angiogenesis. We suggest that an abnormal process of angiogenesis plays a key role in the molecular pathogenesis of BEN.


title: Response surface modeling for hot, humid air decontamination of materials contaminated with Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam spores

text: Response surface methodology using a face-centered cube design was used to describe and predict spore inactivation of Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam spores after exposure of six spore-contaminated materials to hot, humid air. For each strain/material pair, an attempt was made to fit a first or second order model. All three independent predictor variables (temperature, relative humidity, and time) were significant in the models except that time was not significant for B. thuringiensis Al Hakam on nylon. Modeling was unsuccessful for wiring insulation and wet spores because there was complete spore inactivation in the majority of the experimental space. In cases where a predictive equation could be fit, response surface plots with time set to four days were generated. The survival of highly purified Bacillus spores can be predicted for most materials tested when given the settings for temperature, relative humidity, and time. These predictions were cross-checked with spore inactivation measurements.


title: Glycyrrhizic Acid in the Treatment of Liver Diseases: Literature Review

text: Glycyrrhizic acid (GA) is a triterpene glycoside found in the roots of licorice plants (Glycyrrhiza glabra). GA is the most important active ingredient in the licorice root, and possesses a wide range of pharmacological and biological activities. GA coupled with glycyrrhetinic acid and 18-beta-glycyrrhetic acid was developed in China or Japan as an anti-inflammatory, antiviral, and antiallergic drug for liver disease. This review summarizes the current biological activities of GA and its medical applications in liver diseases. The pharmacological actions of GA include inhibition of hepatic apoptosis and necrosis; anti-inflammatory and immune regulatory actions; antiviral effects; and antitumor effects. This paper will be a useful reference for physicians and biologists researching GA and will open the door to novel agents in drug discovery and development from Chinese herbs. With additional research, GA may be more widely used in the treatment of liver diseases or other conditions.


title: Effects and cost of glycyrrhizin in the treatment of upper respiratory tract infections in members of the Japanese maritime self-defense force: Preliminary report of a prospective, randomized, double-blind, controlled, parallel-group, alternate-day treatment assignment clinical trial

text: Background: Upper respiratory tract infections (URTIs) account for at least half of all acute illnesses. Specific antiviral therapy has not been developed against most respiratory viruses thought to cause URTIs. The pharmacologic action of glycyrrhizin has been shown to produce anti-inflammatory activity, modulation of the immune system, inhibition of virus growth, and inactivation of viruses. Objective: The aim of this study was to assess the tolerability, efficacy, and cost of glycyrrhizin in improving the severity and duration of signs and symptoms of URTIs. The primary end point was tolerability, and the secondary and points included improvement in signs and symptoms of URTI and cost. Methods: Members of the Japanese Maritime Self-Defense Force (SDF) treated for URTIs from January 2002 to May 2002 in the SDF Etajima Hospital (Hiroshima, Japan) were eligible for this prospective, randomized, double-blind, controlled, parallel-group, alternate-day treatment assignment study. All patients in this study fulfilled the following enrollment criteria: admitted to the hospital on the first arrival day as an outpatient; fever (body temperature <38.0°C) with signs and symptoms of URTI (headache, sore throat, rhinorrhea, pharyngitis); and had not received antibiotics or oseltamivir phosphate for 4 weeks before the study. Patients who were admitted on an even day received an IV drip infusion of 40 mL of glycyrrhizin (0.2%) and 500 mL of lactated Ringer's solution daily during hospitalization (glycyrrhizin group). Patients who were admitted on an odd day received an IV drip infusion of 500 mL/d of lactated Ringer's solution only (control group). Adverse effects were assessed by the physicians during hospitalization, using patient interview and laboratory analysis. Results: Forty-one consecutive patients entered the study; 15 patients (15 men, 0 women; mean [SD] age, 25.2 [1.5] years) were assigned to the glycyrrhizin group and 269 patients (24 men, 2 women; mean [SD] age, 22.6 [0.9] years) were assigned to the control group. The 2 groups were similar in terms of baseline characteristics. The mean duration of hospitalization was shorter (P = 0.01), the mean maximum body temperature 24 to 48 hours after admission was less (P = 0.05), and the cost of therapy (P = 0.03) was less in the glycyrrhizin group than the control group. No AEs were reported. Conclusions: In this study of hospitalized patients with URTIs, glycyrrhizin therapy was associated with a shorter hospitalization, lower-grade fever, and lower cost of therapy compared with controls, showing that it may be beneficial to patients with URTIs without acute bacterial infections.


title: Circulating Levels of Tumor Necrosis Factor-Alpha Receptor 2 Are Increased in Heart Failure with Preserved Ejection Fraction Relative to Heart Failure with Reduced Ejection Fraction: Evidence for a Divergence in Pathophysiology

text: BACKGROUND: Various pathways have been implicated in the pathogenesis of heart failure (HF) with preserved ejection fraction (HFPEF). Inflammation in response to comorbid conditions, such as hypertension and diabetes, may play a proportionally larger role in HFPEF as compared to HF with reduced ejection fraction (HFREF). METHODS AND RESULTS: This study investigated inflammation mediated by the tumor necrosis factor-alpha (TNFα) axis in community-based cohorts of HFPEF patients (n = 100), HFREF patients (n = 100) and healthy controls (n = 50). Enzyme-linked immunosorbent assays were used to investigate levels of TNFα, its two receptors (TNFR1 and TNFR2), and a non-TNFα cytokine, interleukin-6 (IL-6), in plasma derived from peripheral blood samples. Plasma levels of TNFα and TNFR1 were significantly elevated in HFPEF relative to controls, while levels of TNFR2 were significantly higher in HFPEF than both controls and HFREF. TNFα, TNFR1 and TNFR2 were each significantly associated with at least two of the following: age, estimated glomerular filtration rate, hypertension, diabetes, smoking, peripheral vascular disease or history of atrial fibrillation. TNFR2 levels were also significantly associated with increasing grade of diastolic dysfunction and severity of symptoms in HFPEF. CONCLUSIONS: Inflammation mediated through TNFα and its receptors, TNFR1 and TNFR2, may represent an important component of a comorbidity-induced inflammatory response that partially drives the pathophysiology of HFPEF.


title: Sustained high serum malondialdehyde levels are associated with severity and mortality in septic patients

text: INTRODUCTION: There is a hyperoxidative state in sepsis. The objective of this study was to determine serum malondialdehyde (MDA) levels during the first week of follow up, whether such levels are associated with severity during the first week and whether non-surviving patients showed higher MDA levels than survivors during the first week. METHODS: We performed an observational, prospective, multicenter study in six Spanish Intensive Care Units. Serum levels of MDA were measured in 328 patients (215 survivors and 113 non-survivors) with severe sepsis at days one, four and eight of diagnosis, and in 100 healthy controls. The primary endpoint was 30-day mortality and the secondary endpoint was six -month mortality. The association between continuous variables was carried out using Spearman’s rank correlation coefficient. Cox regression analysis was applied to determine the independent contribution of serum MDA levels on the prediction of 30-day and 6-month mortality. Hazard ratio (HR) and 95% confidence intervals (CI) were calculated as measures of the clinical impact of the predictor variables. RESULTS: We found higher serum MDA in septic patients at day one (p < 0.001), day four (p < 0.001) and day eight (p < 0.001) of diagnosis than in healthy controls. Serum MDA was lower in surviving than non-surviving septic patients at day one (p < 0.001), day four (p < 0.001) and day eight (p < 0.001). Serum MDA levels were positively correlated with lactic acid and SOFA during the first week. Finally, serum MDA levels were associated with 30-day mortality (HR = 1.05; 95% CI = 1.02-1.09; p = 0.005) and six-month mortality (hazard ratio (HR) = 1.05; 95% CI = 1.02-1.09; p = 0.003) after controlling for lactic acid levels, acute physiology and chronic health evaluation (APACHE)-II, diabetes mellitus, bloodstream infection and chronic renal failure. CONCLUSIONS: To our knowledge, this is the largest series providing data on the oxidative state in septic patients to date. The novel finding is that high serum MDA levels sustained throughout the first week of follow up were associated with severity and mortality in septic patients.


title: Kinetics of circulating immunoglobulin M in sepsis: relationship with final outcome

text: INTRODUCTION: The aim of this study was to investigate the kinetics of immunoglobulin M (IgM) during the different stages of sepsis. METHODS: In this prospective multicenter study, blood sampling for IgM measurement was done within the first 24 hours from diagnosis in 332 critically ill patients; in 83 patients this was repeated upon progression to more severe stages. Among these 83 patients, 30 patients with severe sepsis progressed into shock and IgM was monitored daily for seven consecutive days. Peripheral blood mononuclear cells (PBMCs) were isolated from 55 patients and stimulated for IgM production. RESULTS: Serum IgM was decreased in septic shock compared to patients with systemic inflammatory response syndrome (SIRS) and patients with severe sepsis. Paired comparisons at distinct time points of the sepsis course showed that IgM was decreased only when patients deteriorated from severe sepsis to septic shock. Serial measurements in these patients, beginning from the early start of vasopressors, showed that the distribution of IgM over time was significantly greater for survivors than for non-survivors. Production of IgM by PBMCs was significantly lower at all stages of sepsis compared with healthy controls. CONCLUSIONS: Specific changes of circulating IgM occur when patients with severe sepsis progress into septic shock. The distribution of IgM is lower among non-survivors.


Fever management in intensive care patients with infections


title: Inhibition of pulmonary nuclear factor kappa-B decreases the severity of acute Escherichia coli pneumonia but worsens prolonged pneumonia

text: INTRODUCTION: Nuclear factor (NF)-κB is central to the pathogenesis of inflammation in acute lung injury, but also to inflammation resolution and repair. We wished to determine whether overexpression of the NF-κB inhibitor IκBα could modulate the severity of acute and prolonged pneumonia-induced lung injury in a series of prospective randomized animal studies. METHODS: Adult male Sprague-Dawley rats were randomized to undergo intratracheal instillation of (a) 5 × 10(9 )adenoassociated virus (AAV) vectors encoding the IκBα transgene (5 × 10(9 )AAV-IκBα); (b) 1 × 10(10 )AAV-IκBα; (c) 5 × 10(10 )AAV-IκBα; or (d) vehicle alone. After intratracheal inoculation with Escherichia coli, the severity of the lung injury was measured in one series over a 4-hour period (acute pneumonia), and in a second series after 72 hours (prolonged pneumonia). Additional experiments examined the effects of IκBα and null-gene overexpression on E. coli-induced and sham pneumonia. RESULTS: In acute pneumonia, IκBα dose-dependently decreased lung injury, improving arterial oxygenation and lung static compliance, reducing alveolar protein leak and histologic injury, and decreasing alveolar IL-1β concentrations. Benefit was maximal at the intermediate (1 × 10(10)) IκBα vector dose; however, efficacy was diminished at the higher (5 × 10(10)) IκBα vector dose. In contrast, IκBα worsened prolonged pneumonia-induced lung injury, increased lung bacterial load, decreased lung compliance, and delayed resolution of the acute inflammatory response. CONCLUSIONS: Inhibition of pulmonary NF-κB activity reduces early pneumonia-induced injury, but worsens injury and bacterial load during prolonged pneumonia.


title: Patients with community acquired pneumonia admitted to European intensive care units: an epidemiological survey of the GenOSept cohort

text: INTRODUCTION: Community acquired pneumonia (CAP) is the most common infectious reason for admission to the Intensive Care Unit (ICU). The GenOSept study was designed to determine genetic influences on sepsis outcome. Phenotypic data was recorded using a robust clinical database allowing a contemporary analysis of the clinical characteristics, microbiology, outcomes and independent risk factors in patients with severe CAP admitted to ICUs across Europe. METHODS: Kaplan-Meier analysis was used to determine mortality rates. A Cox Proportional Hazards (PH) model was used to identify variables independently associated with 28-day and six-month mortality. RESULTS: Data from 1166 patients admitted to 102 centres across 17 countries was extracted. Median age was 64 years, 62% were male. Mortality rate at 28 days was 17%, rising to 27% at six months. Streptococcus pneumoniae was the commonest organism isolated (28% of cases) with no organism identified in 36%. Independent risk factors associated with an increased risk of death at six months included APACHE II score (hazard ratio, HR, 1.03; confidence interval, CI, 1.01-1.05), bilateral pulmonary infiltrates (HR1.44; CI 1.11-1.87) and ventilator support (HR 3.04; CI 1.64-5.62). Haematocrit, pH and urine volume on day one were all associated with a worse outcome. CONCLUSIONS: The mortality rate in patients with severe CAP admitted to European ICUs was 27% at six months. Streptococcus pneumoniae was the commonest organism isolated. In many cases the infecting organism was not identified. Ventilator support, the presence of diffuse pulmonary infiltrates, lower haematocrit, urine volume and pH on admission were independent predictors of a worse outcome.


title: Plasma kallistatin levels in patients with severe community-acquired pneumonia

text: INTRODUCTION: Community-acquired pneumonia (CAP) requiring intensive care unit (ICU) treatment commonly causes acute respiratory failure with high mortality. Kallistatin, an endogenous tissue kallikrein inhibitor, has been reported to be protective in various human diseases. The aim of this study was to assess the correlations of kallistatin with other biomarkers and to determine whether kallistatin levels have a prognostic value in severe CAP. METHODS: Plasma samples and clinical data were prospectively collected from 54 patients with severe CAP requiring ICU admission. Seventeen healthy control subjects were included for comparison. Plasma kallistatin, kallikrein, and other biomarkers of inflammation (tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, IL-8, C-reactive protein (CRP)), and anti-coagulation (protein C, anti-thrombin III) were measured on days 1 and 4 of ICU admission. Comparison between survivors (n = 41) and nonsurvivors (n = 13) was performed. RESULTS: Plasma kallistatin was significantly consumed in severe CAP patients compared with healthy individuals. Lower day 1 kallistatin levels showed a strong trend toward increased mortality (P = 0.018) and higher day 1 CURB-65 scores (P = 0.004). Plasma kallistatin levels on day 1 of ICU admission were significantly decreased in patients who developed septic shock (P = 0.017) and who had acute respiratory distress syndrome (P = 0.044). In addition, kallistatin levels were positively correlated with anti-thrombin III and protein C and inversely correlated with IL-1β, IL-6, and CRP levels. In a multivariate logistic regression analysis, higher day 1 CURB-65 scores were independent predictors of mortality (odds ratio = 29.9; P = 0.009). Also, higher day 1 kallistatin levels were independently associated with a decreased risk of death (odds ratio, 0.1) with a nearly significant statistical difference (P = 0.056). Furthermore, we found that a cutoff level of 6.5 μg/ml of day 1 kallistatin determined by receiver operating characteristic curves could be used to distinguish between patients who survived in 60 days and those who did not. CONCLUSIONS: These results suggest that kallistatin may serve as a novel marker for severe CAP prognosis and may be involved in the pathogenesis of CAP through antiinflammatory and anticoagulation effects. See related letter by Katz et al., http://ccforum.com/content/17/2/429


title: Extracorporeal membrane oxygenation (ECMO) in patients with H1N1 influenza infection: a systematic review and meta-analysis including 8 studies and 266 patients receiving ECMO

text: INTRODUCTION: H1N1 influenza can cause severe acute lung injury (ALI). Extracorporeal membrane oxygenation (ECMO) can support gas exchange in patients failing conventional mechanical ventilation, but its role is still controversial. We conducted a systematic review and meta-analysis on ECMO for H1N1-associated ALI. METHODS: CENTRAL, Google Scholar, MEDLINE/PubMed and Scopus (updated 2 January 2012) were systematically searched. Studies reporting on 10 or more patients with H1N1 infection treated with ECMO were included. Baseline, procedural, outcome and validity data were systematically appraised and pooled, when appropriate, with random-effect methods. RESULTS: From 1,196 initial citations, 8 studies were selected, including 1,357 patients with confirmed/suspected H1N1 infection requiring intensive care unit admission, 266 (20%) of whom were treated with ECMO. Patients had a median Sequential Organ Failure Assessment (SOFA) score of 9, and had received mechanical ventilation before ECMO implementation for a median of two days. ECMO was implanted before inter-hospital patient transfer in 72% of cases and in most patients (94%) the veno-venous configuration was used. ECMO was maintained for a median of 10 days. Outcomes were highly variable among the included studies, with in-hospital or short-term mortality ranging between 8% and 65%, mainly depending on baseline patient features. Random-effect pooled estimates suggested an overall in-hospital mortality of 28% (95% confidence interval 18% to 37%; I(2 )= 64%). CONCLUSIONS: ECMO is feasible and effective in patients with ALI due to H1N1 infection. Despite this, prolonged support (more than one week) is required in most cases, and subjects with severe comorbidities or multiorgan failure remain at high risk of in-hospital death.


title: Prediction of mortality in adult patients with severe acute lung failure receiving veno-venous extracorporeal membrane oxygenation: a prospective observational study

text: INTRODUCTION: Veno-venous extracorporeal membrane oxygenation (vvECMO) can be a life-saving therapy in patients with severe acute lung failure refractory to conventional therapy. Nevertheless, vvECMO is a procedure associated with high costs and resource utilization. The aim of this study was to assess published models for prediction of mortality following vvECMO and optimize an alternative model. METHODS: Established mortality risk scores were validated to assess their usefulness in 304 adult patients undergoing vvECMO for refractory lung failure at the University Medical Center Regensburg from 2008 to 2013. A parsimonious prediction model was developed based on variables available before ECMO initiation using logistic regression modelling. We then assessed whether addition of variables available one day after ECMO implementation enhanced mortality prediction. Models were internally validated and calibrated by bootstrapping (400 runs). Predictive ability, goodness-of-fit and model discrimination were compared across the different models. RESULTS: In the present study population, existing mortality prediction tools for vvECMO patients showed suboptimal performance. Evaluated before vvECMO initiation, a logistic prediction model comprising age, immunocompromised state, artificial minute ventilation, pre-ECMO serum lactate and hemoglobin concentrations showed best mortality prediction in our patients (area under curve, AUC: 0.75). Additional information about norepinephrine dosage, fraction of inspired oxygen, C-reactive protein and fibrinogen concentrations the first day following ECMO initiation further improved discrimination (AUC: 0.79, P = 0.03) and predictive ability (likelihood ratio test, P < 0.001). When classifying patients as lower (<40%) or higher (>80%) risk based on their predicted mortality, the pre-ECMO and day1-on-ECMO models had negative/positive predictive values of 76%/82% and 82%/81%, respectively. CONCLUSIONS: While pre-ECMO mortality prediction remains a challenge due to large patient heterogeneity, evaluation one day after ECMO initiation may improve the ability to separate lower- and higher-risk patients. Our findings support the clinical perception that chronic health condition, high comorbidity and reduced functional reserves are strongly related to survival during and following ECMO support. Renewed evaluation the first day after ECMO initiation may provide enhanced guidance for further handling of ECMO patients. Despite the usefulness of prediction models, thorough clinical evaluation should always represent the cornerstone in decision for ECMO.


title: Year in review 2012: Critical Care - respiratory infections

text: Over the last two decades, considerable progress has been made in the understanding of disease mechanisms and infection control strategies related to infections, particularly pneumonia, in critically ill patients. Patient-centered and preventative strategies assume paramount importance in this era of limited health-care resources, in which effective targeted therapy is required to achieve the best outcomes. Risk stratification using severity scores and inflammatory biomarkers is a promising strategy for identifying sick patients early during their hospital stay. The emergence of multidrug-resistant pathogens is becoming a major hurdle in intensive care units. Cooperation, education, and interaction between multiple disciplines in the intensive care unit are required to limit the spread of resistant pathogens and to improve care. In this review, we summarize findings from major publications over the last year in the field of respiratory infections in critically ill patients, putting an emphasis on a newer understanding of pathogenesis, use of biomarkers, and antibiotic stewardship and examining new treatment options and preventive strategies.


title: Clinical and biological role of secretory phospholipase A2 in acute respiratory distress syndrome infants

text: INTRODUCTION: Secretory phospholipase A2 is supposed to play a role in acute lung injury but no data are available for pediatric acute respiratory distress syndrome (ARDS). It is not clear which enzyme subtypes are secreted and what the relationships are between enzyme activity, biophysical and biochemical parameters, and clinical outcomes. We aimed to measure the enzyme and identify its subtypes and to study its biochemical and biophysical effect. The secondary aim was to correlate enzyme activity with clinical outcome. METHODS: Bronchoalveolar lavage was performed in 24 infants with ARDS and 14 controls with no lung disease. Samples were assayed for secretory phospholipase A2 and molecules related to its activity and expression. Western blotting and captive bubble surfactometry were also performed. Clinical data were real time downloaded. RESULTS: Tumor necrosis factor-α (814 (506-2,499) vs. 287 (111-1,315) pg/mL; P = 0.04), enzyme activity (430 (253-600) vs. 149 (61-387) IU/mL; P = 0.01), free fatty acids (4.3 (2.8-8.6) vs. 2 (0.8-4.6) mM; P = 0.026), and minimum surface tension (25.6 ± 6.1 vs. 18 ± 1.8 mN/m; P = 0.006) were higher in ARDS than in controls. Phospholipids are lower in ARDS than in controls (76.5 (54-100) vs. 1,094 (536-2,907) μg/mL; P = 0.0001). Three enzyme subtypes were identified (-IIA, -V, -X), although in lower quantities in controls; another subtype (-IB) was mainly detected in ARDS. Significant correlations exist between enzyme activity, free fatty acids (ρ = 0.823; P < 0.001), and surface tension (ρ = 0.55; P < 0.028). Correlations also exist with intensive care stay (ρ = 0.54; P = 0.001), PRISM-III(24 )(ρ = 0.79; P< 0.001), duration of ventilation (ρ = 0.53; P = 0.002), and oxygen therapy (ρ = 0.54; P = 0.001). CONCLUSIONS: Secretory phospholipase A2 activity is raised in pediatric ARDS and constituted of four subtypes. Enzyme correlates with some inflammatory mediators, surface tension, and major clinical outcomes. Secretory phospholipase A2 may be a clinically relevant target in pediatric ARDS.


title: Increased survival of cirrhotic patients with septic shock

text: INTRODUCTION: The overall outcome of septic shock has been recently improved. We sought to determine whether this survival gain extends to the high-risk subgroup of patients with cirrhosis. METHODS: Cirrhotic patients with septic shock admitted to a medical intensive care unit (ICU) during two consecutive periods (1997-2004 and 2005-2010) were retrospectively studied. RESULTS: Forty-seven and 42 cirrhotic patients presented with septic shock in 1997-2004 and 2005-2010, respectively. The recent period differed from the previous one by implementation of adjuvant treatments of septic shock including albumin infusion as fluid volume therapy, low-dose glucocorticoids, and intensive insulin therapy. ICU and hospital survival markedly improved over time (40% in 2005-2010 vs. 17% in 1997-2004, P = 0.02 and 29% in 2005-2010 vs. 6% in 1997-2004, P = 0.009, respectively). Furthermore, this survival gain in the latter period was sustained for 6 months (survival rate 24% in 2005-2010 vs. 6% in 1997-2004, P = 0.06). After adjustment with age, the liver disease stage (Child-Pugh score), and the critical illness severity score (SOFA score), ICU admission between 2005 and 2010 remained an independent favorable prognostic factor (odds ratio (OR) 0.09, 95% confidence interval (CI) 0.02-0.4, P = 0.004). The stage of the underlying liver disease was also independently associated with hospital mortality (Child-Pugh score: OR 1.42 per point, 95% CI 1.06-1.9, P = 0.018). CONCLUSIONS: In the light of advances in management of both cirrhosis and septic shock, survival of such patients substantially increased over recent years. The stage of the underlying liver disease and the related therapeutic options should be included in the decision-making process for ICU admission.


title: Recombinant factor VIIa for uncontrollable bleeding in patients with extracorporeal membrane oxygenation: report on 15 cases and literature review

text: INTRODUCTION: Bleeding is the most frequent complication in patients receiving venoarterial or venovenous extracorporeal membrane oxygenation (ECMO). Recombinant activated factor VII (rFVIIa) has been used in these patients with conflicting results. We describe our experience with rFVIIa for refractory bleeding in this setting and review the cases reported in the literature. METHODS: Clinical characteristics, demographics, bleeding, thrombotic complications, mortality, and rFVIIa administration were retrospectively collected for analysis from the electronic charts of the 15 patients in our intensive care unit who received rFVIIa while being given ECMO from January 2006 to March 2011. RESULTS: Fifteen patients received rFVIIa for persistent bleeding under venoarterial (n = 11) or venovenous (n = 4) ECMO. Bleeding dramatically decreased in 14 patients, without a major thrombotic event, except in one patient in whom a major stroke could not be ruled out. Two circuits were changed within the 48 hours after rFVIIa administration for clots in the membrane and decreased oxygenation but without massive clotting. The mortality rate was 60%. CONCLUSIONS: rFVIIa use for intractable hemorrhaging in patients receiving ECMO controlled bleeding, without major thrombotic events, and with 60% dying. Hence, its use warrants discussion, and clinicians should be aware of the possibility of potentially life-threatening systemic thrombosis, emboli, or circuit clotting. Whether rFVIIa can save the lives of such patients remains to be determined.


title: Extra Corporeal Membrane Oxygenation (ECMO) in three HIV-positive patients with acute respiratory distress syndrome

text: BACKGROUND: Extracorporeal membrane oxygenation (ECMO) is a life-saving bridging procedure in patients with severe acute respiratory distress syndrome (ARDS). Official indications for ECMO are unclear for immunocompromised and HIV-positive patients affected by severe hypoxemia. Uncertainties are related to prognosis and efficacy of treatment of the underlying disease. However, the care of patients with HIV infection has advanced since the introduction of highly active antiretroviral therapy (HAART), with increased life expectancy and decreased mortality. CASE PRESENTATION: Three HIV-infected patients with AIDS were admitted to ICU and were treated with ECMO: a 21 years old Caucasian female with congenital HIV infection presented with Pneumocystis jirovecii pneumonia (PJP); a 38 years old Caucasian female with HIV-HCV infection and L. pneumophila pneumonia; a 24 years old Caucasian male with fever, cough weight loss and PJP pneumonia. Two patients were alive, with a good immunovirological profile and they went back to their previous quality of life. The last patient died with septic shock after three months of ICU stay. CONCLUSION: ECMO was effective in three HIV-positive patients with an otherwise fatal respiratory failure. All patients had severe immunosuppression and/or limited antiretroviral options. A multidisciplinary critical team is needed to individualize the use of ECMO in immunocompromised patients, including those with HIV infection.


title: OutbreakTools: A new platform for disease outbreak analysis using the R software

text: The investigation of infectious disease outbreaks relies on the analysis of increasingly complex and diverse data, which offer new prospects for gaining insights into disease transmission processes and informing public health policies. However, the potential of such data can only be harnessed using a number of different, complementary approaches and tools, and a unified platform for the analysis of disease outbreaks is still lacking. In this paper, we present the new R package OutbreakTools, which aims to provide a basis for outbreak data management and analysis in R. OutbreakTools is developed by a community of epidemiologists, statisticians, modellers and bioinformaticians, and implements classes and methods for storing, handling and visualizing outbreak data. It includes real and simulated outbreak datasets. Together with a number of tools for infectious disease epidemiology recently made available in R, OutbreakTools contributes to the emergence of a new, free and open-source platform for the analysis of disease outbreaks.


title: ELR(+) chemokine signaling in host defense and disease in a viral model of central nervous system disease

text: Intracranial infection of the neurotropic JHM strain of mouse hepatitis virus (JHMV) into the central nervous system (CNS) of susceptible strains of mice results in an acute encephalomyelitis, accompanied by viral replication in glial cells and robust infiltration of virus-specific T cells that contribute to host defense through cytokine secretion and cytolytic activity. Mice surviving the acute stage of disease develop an immune-mediated demyelinating disease, characterized by viral persistence in white matter tracts and a chronic neuroinflammatory response dominated by T cells and macrophages. Chemokines and their corresponding chemokine receptors are dynamically expressed throughout viral infection of the CNS, influencing neuroinflammation by regulating immune cell infltration and glial biology. This review is focused upon the pleiotropic chemokine receptor CXCR2 and its effects upon neutrophils and oligodendrocytes during JHMV infection and a number of other models of CNS inflammation.


title: Flying-Fox Species Density - A Spatial Risk Factor for Hendra Virus Infection in Horses in Eastern Australia

text: Hendra virus causes sporadic but typically fatal infection in horses and humans in eastern Australia. Fruit-bats of the genus Pteropus (commonly known as flying-foxes) are the natural host of the virus, and the putative source of infection in horses; infected horses are the source of human infection. Effective treatment is lacking in both horses and humans, and notwithstanding the recent availability of a vaccine for horses, exposure risk mitigation remains an important infection control strategy. This study sought to inform risk mitigation by identifying spatial and environmental risk factors for equine infection using multiple analytical approaches to investigate the relationship between plausible variables and reported Hendra virus infection in horses. Spatial autocorrelation (Global Moran’s I) showed significant clustering of equine cases at a distance of 40 km, a distance consistent with the foraging ‘footprint’ of a flying-fox roost, suggesting the latter as a biologically plausible basis for the clustering. Getis-Ord Gi* analysis identified multiple equine infection hot spots along the eastern Australia coast from far north Queensland to central New South Wales, with the largest extending for nearly 300 km from southern Queensland to northern New South Wales. Geographically weighted regression (GWR) showed the density of P. alecto and P. conspicillatus to have the strongest positive correlation with equine case locations, suggesting these species are more likely a source of infection of Hendra virus for horses than P. poliocephalus or P. scapulatus. The density of horses, climate variables and vegetation variables were not found to be a significant risk factors, but the residuals from the GWR suggest that additional unidentified risk factors exist at the property level. Further investigations and comparisons between case and control properties are needed to identify these local risk factors.


title: Can’t RIDD off viruses

text: The mammalian genome has evolved to encode a battery of mechanisms, to mitigate a progression in the life cycle of an invasive viral pathogen. Although apparently disadvantaged by their dependence on the host biosynthetic processes, an immensely faster rate of evolution provides viruses with an edge in this conflict. In this review, I have discussed the potential anti-virus activity of inositol-requiring enzyme 1 (IRE1), a well characterized effector of the cellular homeostatic response to an overloading of the endoplasmic reticulum (ER) protein-folding capacity. IRE1, an ER-membrane-resident ribonuclease (RNase), upon activation catalyses regulated cleavage of select protein-coding and non-coding host RNAs, using an RNase domain which is homologous to that of the known anti-viral effector RNaseL. The latter operates as part of the Oligoadenylate synthetase OAS/RNaseL system of anti-viral defense mechanism. Protein-coding RNA substrates are differentially treated by the IRE1 RNase to either augment, through cytoplasmic splicing of an intron in the Xbp1 transcript, or suppress gene expression. This referred suppression of gene expression is mediated through degradative cleavage of a select cohort of cellular RNA transcripts, initiating the regulated IRE1-dependent decay (RIDD) pathway. The review first discusses the anti-viral mechanism of the OAS/RNaseL system and evasion tactics employed by different viruses. This is followed by a review of the RIDD pathway and its potential effect on the stability of viral RNAs. I conclude with a comparison of the enzymatic activity of the two RNases followed by deliberations on the physiological consequences of their activation.


title: A fast and robust iterative algorithm for prediction of RNA pseudoknotted secondary structures

text: BACKGROUND: Improving accuracy and efficiency of computational methods that predict pseudoknotted RNA secondary structures is an ongoing challenge. Existing methods based on free energy minimization tend to be very slow and are limited in the types of pseudoknots that they can predict. Incorporating known structural information can improve prediction accuracy; however, there are not many methods for prediction of pseudoknotted structures that can incorporate structural information as input. There is even less understanding of the relative robustness of these methods with respect to partial information. RESULTS: We present a new method, Iterative HFold, for pseudoknotted RNA secondary structure prediction. Iterative HFold takes as input a pseudoknot-free structure, and produces a possibly pseudoknotted structure whose energy is at least as low as that of any (density-2) pseudoknotted structure containing the input structure. Iterative HFold leverages strengths of earlier methods, namely the fast running time of HFold, a method that is based on the hierarchical folding hypothesis, and the energy parameters of HotKnots V2.0. Our experimental evaluation on a large data set shows that Iterative HFold is robust with respect to partial information, with average accuracy on pseudoknotted structures steadily increasing from roughly 54% to 79% as the user provides up to 40% of the input structure. Iterative HFold is much faster than HotKnots V2.0, while having comparable accuracy. Iterative HFold also has significantly better accuracy than IPknot on our HK-PK and IP-pk168 data sets. CONCLUSIONS: Iterative HFold is a robust method for prediction of pseudoknotted RNA secondary structures, whose accuracy with more than 5% information about true pseudoknot-free structures is better than that of IPknot, and with about 35% information about true pseudoknot-free structures compares well with that of HotKnots V2.0 while being significantly faster. Iterative HFold and all data used in this work are freely available at http://www.cs.ubc.ca/~hjabbari/software.php.


title: Characterization of the Ectodomain of the Envelope Protein of Dengue Virus Type 4: Expression, Membrane Association, Secretion and Particle Formation in the Absence of Precursor Membrane Protein

text: BACKGROUND: The envelope (E) of dengue virus (DENV) is the major target of neutralizing antibodies and vaccine development. After biosynthesis E protein forms a heterodimer with precursor membrane (prM) protein. Recent reports of infection enhancement by anti-prM monoclonal antibodies (mAbs) suggest anti-prM responses could be potentially harmful. Previously, we studied a series of C-terminal truncation constructs expressing DENV type 4 prM/E or E proteins and found the ectodomain of E protein alone could be recognized by all 12 mAbs tested, suggesting E protein ectodomain as a potential subunit immunogen without inducing anti-prM response. The characteristics of DENV E protein ectodomain in the absence of prM protein remains largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the expression, membrane association, glycosylation pattern, secretion and particle formation of E protein ectodomain of DENV4 in the presence or absence of prM protein. E protein ectodomain associated with membrane in or beyond trans-Golgi and contained primarily complex glycans, whereas full-length E protein associated with ER membrane and contained high mannose glycans. In the absence of prM protein, E protein ectodomain can secrete as well as form particles of approximately 49 nm in diameter, as revealed by sucrose gradient ultracentrifugation with or without detergent and electron microscopy. Mutational analysis revealed that the secretion of E protein ectodomain was affected by N-linked glycosylation and could be restored by treatment with ammonia chloride. CONCLUSIONS/SIGNIFICANCE: Considering the enhancement of DENV infectivity by anti-prM antibodies, our findings provide new insights into the expression and secretion of E protein ectodomain in the absence of prM protein and contribute to future subunit vaccine design.


title: Analysis of tetra- and hepta-nucleotides motifs promoting -1 ribosomal frameshifting in Escherichia coli

text: Programmed ribosomal -1 frameshifting is a non-standard decoding process occurring when ribosomes encounter a signal embedded in the mRNA of certain eukaryotic and prokaryotic genes. This signal has a mandatory component, the frameshift motif: it is either a Z_ZZN tetramer or a X_XXZ_ZZN heptamer (where ZZZ and XXX are three identical nucleotides) allowing cognate or near-cognate repairing to the -1 frame of the A site or A and P sites tRNAs. Depending on the signal, the frameshifting frequency can vary over a wide range, from less than 1% to more than 50%. The present study combines experimental and bioinformatics approaches to carry out (i) a systematic analysis of the frameshift propensity of all possible motifs (16 Z_ZZN tetramers and 64 X_XXZ_ZZN heptamers) in Escherichia coli and (ii) the identification of genes potentially using this mode of expression amongst 36 Enterobacteriaceae genomes. While motif efficiency varies widely, a major distinctive rule of bacterial -1 frameshifting is that the most efficient motifs are those allowing cognate re-pairing of the A site tRNA from ZZN to ZZZ. The outcome of the genomic search is a set of 69 gene clusters, 59 of which constitute new candidates for functional utilization of -1 frameshifting.


title: Results From the First Six Years of National Sentinel Surveillance for Influenza in Kenya, July 2007–June 2013

text: BACKGROUND: Recent studies have shown that influenza is associated with significant disease burden in many countries in the tropics, but until recently national surveillance for influenza was not conducted in most countries in Africa. METHODS: In 2007, the Kenyan Ministry of Health with technical support from the CDC-Kenya established a national sentinel surveillance system for influenza. At 11 hospitals, for every hospitalized patient with severe acute respiratory illness (SARI), and for the first three outpatients with influenza-like illness (ILI) per day, we collected both nasopharyngeal and oropharyngeal swabs. Beginning in 2008, we conducted in-hospital follow-up for SARI patients to determine outcome. Specimens were tested by real time RT-PCR for influenza A and B. Influenza A-positive specimens were subtyped for H1, H3, H5, and (beginning in May 2009) A(H1N1)pdm09. RESULTS: From July 1, 2007 through June 30, 2013, we collected specimens from 24,762 SARI and 14,013 ILI patients. For SARI and ILI case-patients, the median ages were 12 months and 16 months, respectively, and 44% and 47% were female. In all, 2,378 (9.6%) SARI cases and 2,041 (14.6%) ILI cases were positive for influenza viruses. Most influenza-associated SARI cases (58.6%) were in children <2 years old. Of all influenza-positive specimens, 78% were influenza A, 21% were influenza B, and 1% were influenza A/B coinfections. Influenza circulated in every month. In four of the six years influenza activity peaked during July–November. Of 9,419 SARI patients, 2.7% died; the median length of hospitalization was 4 days. CONCLUSIONS: During six years of surveillance in Kenya, influenza was associated with nearly 10 percent of hospitalized SARI cases and one-sixth of outpatient ILI cases. Most influenza-associated SARI and ILI cases were in children <2 years old; interventions to reduce the burden of influenza, such as vaccine, could consider young children as a priority group.


title: From gene to protein—experimental and clinical studies of ACE2 in blood pressure control and arterial hypertension

text: Hypertension is a major risk factor for stroke, coronary events, heart and renal failure, and the renin-angiotensin system (RAS) plays a major role in its pathogenesis. Within the RAS, angiotensin converting enzyme (ACE) converts angiotensin (Ang) I into the vasoconstrictor Ang II. An “alternate” arm of the RAS now exists in which ACE2 counterbalances the effects of the classic RAS through degradation of Ang II, and generation of the vasodilator Ang 1-7. ACE2 is highly expressed in the heart, blood vessels, and kidney. The catalytically active ectodomain of ACE2 undergoes shedding, resulting in ACE2 in the circulation. The ACE2 gene maps to a quantitative trait locus on the X chromosome in three strains of genetically hypertensive rats, suggesting that ACE2 may be a candidate gene for hypertension. It is hypothesized that disruption of tissue ACE/ACE2 balance results in changes in blood pressure, with increased ACE2 expression protecting against increased blood pressure, and ACE2 deficiency contributing to hypertension. Experimental hypertension studies have measured ACE2 in either the heart or kidney and/or plasma, and have reported that deletion or inhibition of ACE2 leads to hypertension, whilst enhancing ACE2 protects against the development of hypertension, hence increasing ACE2 may be a therapeutic option for the management of high blood pressure in man. There have been relatively few studies of ACE2, either at the gene or the circulating level in patients with hypertension. Plasma ACE2 activity is low in healthy subjects, but elevated in patients with cardiovascular risk factors or cardiovascular disease. Genetic studies have investigated ACE2 gene polymorphisms with either hypertension or blood pressure, and have produced largely inconsistent findings. This review discusses the evidence regarding ACE2 in experimental hypertension models and the association between circulating ACE2 activity and ACE2 polymorphisms with blood pressure and arterial hypertension in man.


title: Standards for Sequencing Viral Genomes in the Era of High-Throughput Sequencing

text: Thanks to high-throughput sequencing technologies, genome sequencing has become a common component in nearly all aspects of viral research; thus, we are experiencing an explosion in both the number of available genome sequences and the number of institutions producing such data. However, there are currently no common standards used to convey the quality, and therefore utility, of these various genome sequences. Here, we propose five “standard” categories that encompass all stages of viral genome finishing, and we define them using simple criteria that are agnostic to the technology used for sequencing. We also provide genome finishing recommendations for various downstream applications, keeping in mind the cost-benefit trade-offs associated with different levels of finishing. Our goal is to define a common vocabulary that will allow comparison of genome quality across different research groups, sequencing platforms, and assembly techniques.


title: Plasmodium vivax Antigen Discovery Based on Alpha-Helical Coiled Coil Protein Motif

text: Protein α-helical coiled coil structures that elicit antibody responses, which block critical functions of medically important microorganisms, represent a means for vaccine development. By using bioinformatics algorithms, a total of 50 antigens with α-helical coiled coil motifs orthologous to Plasmodium falciparum were identified in the P. vivax genome. The peptides identified in silico were chemically synthesized; circular dichroism studies indicated partial or high α-helical content. Antigenicity was evaluated using human sera samples from malaria-endemic areas of Colombia and Papua New Guinea. Eight of these fragments were selected and used to assess immunogenicity in BALB/c mice. ELISA assays indicated strong reactivity of serum samples from individuals residing in malaria-endemic regions and sera of immunized mice, with the α-helical coiled coil structures. In addition, ex vivo production of IFN-γ by murine mononuclear cells confirmed the immunogenicity of these structures and the presence of T-cell epitopes in the peptide sequences. Moreover, sera of mice immunized with four of the eight antigens recognized native proteins on blood-stage P. vivax parasites, and antigenic cross-reactivity with three of the peptides was observed when reacted with both the P. falciparum orthologous fragments and whole parasites. Results here point to the α-helical coiled coil peptides as possible P. vivax malaria vaccine candidates as were observed for P. falciparum. Fragments selected here warrant further study in humans and non-human primate models to assess their protective efficacy as single components or assembled as hybrid linear epitopes.


title: Epidemiology and Viral Etiology of the Influenza-Like Illness in Corsica during the 2012–2013 Winter: An Analysis of Several Sentinel Surveillance Systems

text: Influenza-like illness (ILI) surveillance is important to identify circulating and emerging/reemerging strains and unusual epidemiological trends. The present study aimed to give an accurate picture of the 2012–2013 ILI outbreak in Corsica by combining data from several surveillance systems: general practice, emergency general practice, hospital emergency units, intensive care units, and nursing homes. Twenty-eight respiratory viruses were retrospectively investigated from patients in general practice with ILI. Sequence analysis of the genetic changes in the hemagglutinin gene of influenza viruses (A(H1N1)pdm2009, A(H3N2) and B) was performed. The trends in ILI/influenza consultation rates and the relative illness ratios (RIRs) of having an ILI consultation were estimated by age group for the different surveillance systems analyzed. Of the 182 ILI patients enrolled by general practitioners, 57.7% tested positive for influenza viruses. Phylogenetic analyses suggested a genetic drift for influenza B and A(H3N2) viruses. The ILI/influenza surveillance systems showed similar trends and were well correlated. In accordance with virological data, the RIRs of having an ILI consultation were highest among the young (<15 years old) and decreased with age. No clusters of acute respiratory illness were declared by the sentinel nursing homes. This study is noteworthy in that it is the first extensive description of the 2012–2013 ILI outbreak in Corsica as monitored through several surveillance systems. To improve ILI surveillance in Corsica, a consortium that links together the complementary regional surveillance ILI systems described here is being implemented.


title: A Multi-Method Approach to Curriculum Development for In-Service Training in China’s Newly Established Health Emergency Response Offices

text: OBJECTIVE: To describe an innovative approach for developing and implementing an in-service curriculum in China for staff of the newly established health emergency response offices (HEROs), and that is generalisable to other settings. METHODS: The multi-method training needs assessment included reviews of the competency domains needed to implement the International Health Regulations (2005) as well as China’s policies and emergency regulations. The review, iterative interviews and workshops with experts in government, academia, the military, and with HERO staff were reviewed critically by an expert technical advisory panel. FINDINGS: Over 1600 participants contributed to curriculum development. Of the 18 competency domains identified as essential for HERO staff, nine were developed into priority in-service training modules to be conducted over 2.5 weeks. Experts from academia and experienced practitioners prepared and delivered each module through lectures followed by interactive problem-solving exercises and desktop simulations to help trainees apply, experiment with, and consolidate newly acquired knowledge and skills. CONCLUSION: This study adds to the emerging literature on China’s enduring efforts to strengthen its emergency response capabilities since the outbreak of SARS in 2003. The multi-method approach to curriculum development in partnership with senior policy-makers, researchers, and experienced practitioners can be applied in other settings to ensure training is responsive and customized to local needs, resources and priorities. Ongoing curriculum development should reflect international standards and be coupled with the development of appropriate performance support systems at the workplace for motivating staff to apply their newly acquired knowledge and skills effectively and creatively.


title: Discovery of Novel GPVI Receptor Antagonists by Structure-Based Repurposing

text: Inappropriate platelet aggregation creates a cardiovascular risk that is largely managed with thienopyridines and aspirin. Although effective, these drugs carry risks of increased bleeding and drug ‘resistance’, underpinning a drive for new antiplatelet agents. To discover such drugs, one strategy is to identify a suitable druggable target and then find small molecules that modulate it. A good and unexploited target is the platelet collagen receptor, GPVI, which promotes thrombus formation. To identify inhibitors of GPVI that are safe and bioavailable, we docked a FDA-approved drug library into the GPVI collagen-binding site in silico. We now report that losartan and cinanserin inhibit GPVI-mediated platelet activation in a selective, competitive and dose-dependent manner. This mechanism of action likely underpins the cardioprotective effects of losartan that could not be ascribed to its antihypertensive effects. We have, therefore, identified small molecule inhibitors of GPVI-mediated platelet activation, and also demonstrated the utility of structure-based repurposing.


title: HIV-1 and Its gp120 Inhibits the Influenza A(H1N1)pdm09 Life Cycle in an IFITM3-Dependent Fashion

text: HIV-1-infected patients co-infected with A(H1N1)pdm09 surprisingly presented benign clinical outcome. The knowledge that HIV-1 changes the host homeostatic equilibrium, which may favor the patient resistance to some co-pathogens, prompted us to investigate whether HIV-1 infection could influence A(H1N1)pdm09 life cycle in vitro. We show here that exposure of A(H1N1)pdm09-infected epithelial cells to HIV-1 viral particles or its gp120 enhanced by 25% the IFITM3 content, resulting in a decrease in influenza replication. This event was dependent on toll-like receptor 2 and 4. Moreover, knockdown of IFITM3 prevented HIV-1 ability to inhibit A(H1N1)pdm09 replication. HIV-1 infection also increased IFITM3 levels in human primary macrophages by almost 100%. Consequently, the arrival of influenza ribonucleoproteins (RNPs) to nucleus of macrophages was inhibited, as evaluated by different approaches. Reduction of influenza RNPs entry into the nucleus tolled A(H1N1)pdm09 life cycle in macrophages earlier than usual, limiting influenza's ability to induce TNF-α. As judged by analysis of the influenza hemagglutin (HA) gene from in vitro experiments and from samples of HIV-1/A(H1N1)pdm09 co-infected individuals, the HIV-1-induced reduction of influenza replication resulted in delayed viral evolution. Our results may provide insights on the mechanisms that may have attenuated the clinical course of Influenza in HIV-1/A(H1N1)pdm09 co-infected patients during the recent influenza form 2009/2010.


title: A Comprehensive Phylogenetic and Structural Analysis of the Carcinoembryonic Antigen (CEA) Gene Family

text: The carcinoembryonic antigen (CEA) gene family belongs to the immunoglobulin (Ig) superfamily and codes for a vast number of glycoproteins that differ greatly both in amino acid composition and function. The CEA family is divided into two groups, the carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) and the pregnancy-specific glycoproteins. The CEA family members are implicated in pleiotropic (patho)physiological functions including cell–cell adhesion, pregnancy, immunity, neovascularization, regulation of insulin homeostasis, and carcinogenesis. In general, the CEA-encoded proteins are composed of an extracellular region with Ig variable and constant-like domains and a cytoplasmic region containing signaling motifs. Of particular interest, the well-studied human and mouse CEA genes are arranged in clusters in a single chromosome. Taking into account this characteristic, we made an effort to reconstruct the evolutionary history of the CEA gene family. Toward this end, the publicly available genomes were searched extensively for CEA homologs. The domain organization of the retrieved protein sequences was analyzed, and, subsequently, comprehensive phylogenetic analyses of the entire length CEA homologous proteins were performed. A series of evolutionarily conserved amino acid residues, functionally important, were identified. The relative positioning of these residues on the modeled tertiary structure of novel CEA protein domains revealed that they are, also, spatially conserved. Furthermore, the chromosomal arrangement of CEA genes was examined, and it was found that the CEA genes are preserved in terms of position, transcriptional orientation, and number in all species under investigation.


title: Effects of flavonoid-induced oxidative stress on anti-H5N1 influenza a virus activity exerted by baicalein and biochanin A

text: BACKGROUND: Different flavonoids are known to interfere with influenza A virus replication. Recently, we showed that the structurally similar flavonoids baicalein and biochanin A inhibit highly pathogenic avian H5N1 influenza A virus replication by different mechanisms in A549 lung cells. Here, we investigated the effects of both compounds on H5N1-induced reactive oxygen species (ROS) formation and the role of ROS formation during H5N1 replication. FINDINGS: Baicalein and biochanin A enhanced H5N1-induced ROS formation in A549 cells and primary human monocyte-derived macrophages. Suppression of ROS formation induced by baicalein and biochanin A using the antioxidant N-acetyl-L-cysteine strongly increased the anti-H5N1 activity of both compounds in A549 cells but not in macrophages. CONCLUSIONS: These findings emphasise that flavonoids induce complex pharmacological actions some of which may interfere with H5N1 replication while others may support H5N1 replication. A more detailed understanding of these actions and the underlying structure-activity relationships is needed to design agents with optimised anti-H5N1 activity.


title: Advances in Diagnosis of Respiratory Diseases of Small Ruminants

text: Irrespective of aetiology, infectious respiratory diseases of sheep and goats contribute to 5.6 percent of the total diseases of small ruminants. These infectious respiratory disorders are divided into two groups: the diseases of upper respiratory tract, namely, nasal myiasis and enzootic nasal tumors, and diseases of lower respiratory tract, namely, peste des petits ruminants (PPR), parainfluenza, Pasteurellosis, Ovine progressive pneumonia, mycoplasmosis, caprine arthritis encephalitis virus, caseous lymphadenitis, verminous pneumonia, and many others. Depending upon aetiology, many of them are acute and fatal in nature. Early, rapid, and specific diagnosis of such diseases holds great importance to reduce the losses. The advanced enzyme-linked immunosorbent assays (ELISAs) for the detection of antigen as well as antibodies directly from the samples and molecular diagnostic assays along with microsatellites comprehensively assist in diagnosis as well as treatment and epidemiological studies. The present review discusses the advancements made in the diagnosis of common infectious respiratory diseases of sheep and goats. It would update the knowledge and help in adapting and implementing appropriate, timely, and confirmatory diagnostic procedures. Moreover, it would assist in designing appropriate prevention protocols and devising suitable control strategies to overcome respiratory diseases and alleviate the economic losses.


title: Immunogenicity and efficacy of a plasmid DNA rabies vaccine incorporating Myd88 as a genetic adjuvant

text: PURPOSE: Myeloid differentiation factor 88 (Myd88), a ubiquitous Toll-like receptor adaptor molecule, has been reported to play important roles in B cell responses to infections and vaccination. The present study evaluated the effects of genetic adjuvanting with Myd88 on the immune responses to a plasmid DNA rabies vaccine. MATERIALS AND METHODS: Plasmids encoding rabies glycoprotein alone (pIRES-Rgp) or a fragment of Myd88 gene in addition (pIRES-Rgp-Myd) were constructed and administered intramuscularly or intrademally in Swiss albino mice (on days 0, 7, and 21). Rabies virus neutralizing antibody (RVNA) titres were estimated in the mice sera on days 14 and 28 by rapid fluorescent focus inhibition test. The protective efficacy of the constructs was evaluated by an intracerebral challenge with challenge virus standard virus on day 35. RESULTS: Co-expression of Myd88 increased RVNA responses to pIRES-Rgp by 3- and 2-folds, following intramuscular and intradermal immunization, respectively. pIRES-Rgp protected 80% of the mice following intramuscular and intradermal immunizations, while pIRES-Rgp-Myd afforded 100% protection following similar administrations. CONCLUSION: Genetic adjuvanting with Myd88 enhanced the RVNA responses and protective efficacy of a plasmid DNA rabies vaccine. This strategy might be useful for rabies vaccination of canines in the field, and needs further evaluation.


title: Outcomes of Influenza A(H1N1)pdm09 Virus Infection: Results from Two International Cohort Studies

text: BACKGROUND: Data from prospectively planned cohort studies on risk of major clinical outcomes and prognostic factors for patients with influenza A(H1N1)pdm09 virus are limited. In 2009, in order to assess outcomes and evaluate risk factors for progression of illness, two cohort studies were initiated: FLU 002 in outpatients and FLU 003 in hospitalized patients. METHODS AND FINDINGS: Between October 2009 and December 2012, adults with influenza-like illness (ILI) were enrolled; outpatients were followed for 14 days and inpatients for 60 days. Disease progression was defined as hospitalization and/or death for outpatients, and hospitalization for >28 days, transfer to intensive care unit (ICU) if enrolled from general ward, and/or death for inpatients. Infection was confirmed by RT-PCR. 590 FLU 002 and 392 FLU 003 patients with influenza A (H1N1)pdm09 were enrolled from 81 sites in 17 countries at 2 days (IQR 1–3) and 6 days (IQR 4–10) following ILI onset, respectively. Disease progression was experienced by 29 (1 death) outpatients (5.1%; 95% CI: 3.4–7.2%) and 80 inpatients [death (32), hospitalization >28 days (43) or ICU transfer (20)] (21.6%; 95% CI: 17.5–26.2%). Disease progression (death) for hospitalized patients was 53.1% (26.6%) and 12.8% (3.8%), respectively, for those enrolled in the ICU and general ward. In pooled analyses for both studies, predictors of disease progression were age, longer duration of symptoms at enrollment and immunosuppression. Patients hospitalized during the pandemic period had a poorer prognosis than in subsequent seasons. CONCLUSIONS: Patients with influenza A(H1N1)pdm09, particularly when requiring hospital admission, are at high risk for disease progression, especially if they are older, immunodeficient, or admitted late in infection. These data reinforce the need for international trials of novel treatment strategies for influenza infection and serve as a reminder of the need to monitor the severity of seasonal and pandemic influenza epidemics globally. TRIAL REGISTRATION: ClinicalTrials.gov Identifiers: FLU 002- NCT01056354, FLU 003- NCT01056185.


title: Stearoyl-CoA desaturase inhibition blocks formation of hepatitis C virus-induced specialized membranes

text: Hepatitis C virus (HCV) replication is dependent on the formation of specialized membrane structures; however, the host factor requirements for the formation of these HCV complexes remain unclear. Herein, we demonstrate that inhibition of stearoyl-CoA desaturase 1 (SCD-1) halts the biosynthesis of unsaturated fatty acids, such as oleic acid, and negatively modulates HCV replication. Unsaturated fatty acids play key roles in membrane curvature and fluidity. Mechanistically, we demonstrate that SCD-1 inhibition disrupts the integrity of membranous HCV replication complexes and renders HCV RNA susceptible to nuclease-mediated degradation. Our work establishes a novel function for unsaturated fatty acids in HCV replication.


title: Construction of fat1 Gene Expression Vector and Its Catalysis Efficiency in Bovine Fetal Fibroblast Cells

text: The FAT-1 protein is an n-3 fatty acid desaturase, which can recognize a range of 18- and 20-carbon n-6 substrates and transform n-6 polyunsaturated fatty acids (PUFAs) into n-3 PUFAs while n-3 PUFAs have beneficial effect on human health. Fat1 gene is the coding sequence from Caenorhabditis elegans which might play an important role on lipometabolism. To reveal the function of fat1 gene in bovine fetal fibroblast cells and gain the best cell nuclear donor for transgenic bovines, the codon of fat1 sequence was optimized based on the codon usage frequency preference of bovine muscle protein, and directionally cloned into the eukaryotic expression vector pEF-GFP. After identifying by restrictive enzyme digests with AatII/XbaI and sequencing, the fusion plasmid pEF-GFP-fat1 was identified successfully. The pEF-GFP-fat1 vector was transfected into bovine fetal fibroblast cells mediated by Lipofectamine2000(TM). The positive bovine fetal fibroblast cells were selected by G418 and detected by RT-PCR. The results showed that a 1,234 bp transcription was amplified by reverse transcription PCR and the positive transgenic fat1 cell line was successfully established. Then the expression level of fat1 gene in positive cells was detected using quantitative PCR, and the catalysis efficiency was detected by gas chromatography. The results demonstrated that the catalysis efficiency of fat1 was significantly high, which can improve the total PUFAs rich in EPA, DHA and DPA. Construction and expression of pEF-GFP-fat1 vector should be helpful for further understanding the mechanism of regulation of fat1 in vitro. It could also be the first step in the production of fat1 transgenic cattle.


title: Selection of Reference Genes for Gene Expression Studies in Porcine Whole Blood and Peripheral Blood Mononuclear Cells under Polyinosinic:Polycytidylic Acid Stimulation

text: Investigating gene expression of immune cells of whole blood or peripheral blood mononuclear cells (PBMC) under polyinosinic:polycytidylic acid (poly I:C) stimulation is valuable for understanding the immune response of organism to RNA viruses. Quantitative real-time PCR (qRT-PCR) is a standard method for quantification of gene expression studies. However, the reliability of qRT-PCR data critically depends on proper selection of reference genes. In the study, using two different analysis programs, geNorm and NormFinder, we systematically evaluated the gene expression stability of six candidate reference genes (GAPDH, ACTB, B2M, RPL4, TBP, and PPIA) in samples of whole blood and PBMC with or without poly I:C stimulation. Generally, the six candidate genes performed a similar trend of expression stability in the samples of whole blood and PBMC, but more stably expressed in whole blood than in PBMC. geNorm ranked B2M and PPIA as the best combination for gene expression normalization, while according to NormFinder, TBP was ranked as the most stable reference gene, followed by B2M and PPIA. Comprehensively considering the results from the two programs, we recommended using the geometric mean of the three genes, TBP, PPIA and B2M, to normalize the gene expression of whole blood and PBMC with poly I:C stimulation. Our study is the first detailed survey of the gene expression stability in whole blood and PBMC with or without poly I:C stimulation and should be helpful for investigating the molecular mechanism involved in porcine whole blood and PBMC in response to poly I:C stimulation.


title: ICoVax 2013: The 3(rd )ISV Pre-conference Computational Vaccinology Workshop

text: Following last year's computational vaccinology workshop in Shanghai, China, the third ISV Pre-conference Computational Vaccinology Workshop (ICoVax 2013) was held in Barcelona, Spain. ICoVax 2013 provided an international platform for the attendees to showcase their research and discuss problems and solutions in the development and application of computational vaccinology and vaccine informatics tools. The first of the three full-length papers presented at ICoVax discussed the discovery of viral "camouflage" through cross-conservation of T-cell epitopes using a tool called JanusMatrix. This important paper reports that viruses may camouflage their presence in the human body by incorporating sequences in their proteins that are highly cross-conserved at the T-cell receptor surface with human genome proteins, a discovery that has wide ranging implications for the development of vaccines against viruses that use the camouflage method. The other papers described a database for storing experimentally verified data on DNA vaccines and compared therapeutic targets of western drugs to Chinese herbal medicines for cardiovascular diseases. The short poster presentations covered various uses of informatics tools for processing the DNA and microRNA of pathogens to improve vaccine coverage, efficacy and development. A live (on-line) demonstration of the vaccine design toolkit, iVax, presented by Frances Terry of EpiVax, illustrated how computational vaccinology could be used in the design of next generation vaccines.


title: Capacity building efforts and perceptions for wildlife surveillance to detect zoonotic pathogens: comparing stakeholder perspectives

text: BACKGROUND: The capacity to conduct zoonotic pathogen surveillance in wildlife is critical for the recognition and identification of emerging health threats. The PREDICT project, a component of United States Agency for International Development’s Emerging Pandemic Threats program, has introduced capacity building efforts to increase zoonotic pathogen surveillance in wildlife in global ‘hot spot’ regions where zoonotic disease emergence is likely to occur. Understanding priorities, challenges, and opportunities from the perspectives of the stakeholders is a key component of any successful capacity building program. METHODS: A survey was administered to wildlife officials and to PREDICT-implementing in-country project scientists in 16 participating countries in order to identify similarities and differences in perspectives between the groups regarding capacity needs for zoonotic pathogen surveillance in wildlife. RESULTS: Both stakeholder groups identified some human-animal interfaces (i.e. areas of high contact between wildlife and humans with the potential risk for disease transmission), such as hunting and markets, as important for ongoing targeting of wildlife surveillance. Similarly, findings regarding challenges across stakeholder groups showed some agreement in that a lack of sustainable funding across regions was the greatest challenge for conducting wildlife surveillance for zoonotic pathogens (wildlife officials: 96% and project scientists: 81%). However, the opportunity for improving zoonotic pathogen surveillance capacity identified most frequently by wildlife officials as important was increasing communication or coordination among agencies, sectors, or regions (100% of wildlife officials), whereas the most frequent opportunities identified as important by project scientists were increasing human capacity, increasing laboratory capacity, and the growing interest or awareness regarding wildlife disease or surveillance programs (all identified by 69% of project scientists). CONCLUSIONS: A One Health approach to capacity building applied at local and global scales will have the greatest impact on improving zoonotic pathogen surveillance in wildlife. This approach will involve increasing communication and cooperation across ministries and sectors so that experts and stakeholders work together to identify and mitigate surveillance gaps. Over time, this transdisciplinary approach to capacity building will help overcome existing challenges and promote efficient targeting of high risk interfaces for zoonotic pathogen transmission.


Closing the gap: the challenges in converging theoretical, computational, experimental and real-life studies in virus evolution


title: Multiplex Evaluation of Influenza Neutralizing Antibodies with Potential Applicability to In-Field Serological Studies

text: The increased number of outbreaks of H5 and H7 LPAI and HPAI viruses in poultry has major public and animal health implications. The continuous rapid evolution of these subtypes and the emergence of new variants influence the ability to undertake effective surveillance. Retroviral pseudotypes bearing influenza haemagglutinin (HA) and neuraminidase (NA) envelope glycoproteins represent a flexible platform for sensitive, readily standardized influenza serological assays. We describe a multiplex assay for the study of neutralizing antibodies that are directed against both influenza H5 and H7 HA. This assay permits the measurement of neutralizing antibody responses against two antigenically distinct HAs in the same serum/plasma sample thus increasing the amount and quality of serological data that can be acquired from valuable sera. Sera obtained from chickens vaccinated with a monovalent H5N2 vaccine, chickens vaccinated with a bivalent H7N1/H5N9 vaccine, or turkeys naturally infected with an H7N3 virus were evaluated in this assay and the results correlated strongly with data obtained by HI assay. We show that pseudotypes are highly stable under basic cold-chain storage conditions and following multiple rounds of freeze-thaw. We propose that this robust assay may have practical utility for in-field serosurveillance and vaccine studies in resource-limited regions worldwide.


title: Validation of three geolocation strategies for health-facility attendees for research and public health surveillance in a rural setting in western Kenya

text: Understanding the spatial distribution of disease is critical for effective disease control. Where formal address networks do not exist, tracking spatial patterns of clinical disease is difficult. Geolocation strategies were tested at rural health facilities in western Kenya. Methods included geocoding residence by head of compound, participatory mapping and recording the self-reported nearest landmark. Geocoding was able to locate 72·9% [95% confidence interval (CI) 67·7–77·6] of individuals to within 250 m of the true compound location. The participatory mapping exercise was able to correctly locate 82·0% of compounds (95% CI 78·9–84·8) to a 2 × 2·5 km area with a 500 m buffer. The self-reported nearest landmark was able to locate 78·1% (95% CI 73·8–82·1) of compounds to the correct catchment area. These strategies tested provide options for quickly obtaining spatial information on individuals presenting at health facilities.


title: Synergistic Up-Regulation of CXCL10 by Virus and IFN γ in Human Airway Epithelial Cells

text: Airway epithelial cells are the first line of defense against viral infections and are instrumental in coordinating the inflammatory response. In this study, we demonstrate the synergistic stimulation of CXCL10 mRNA and protein, a key chemokine responsible for the early immune response to viral infection, following treatment of airway epithelial cells with IFN γ and influenza virus. The synergism also occurred when the cells were treated with IFN γ and a viral replication mimicker (dsRNA) both in vitro and in vivo. Despite the requirement of type I interferon (IFNAR) signaling in dsRNA-induced CXCL10, the synergism was independent of the IFNAR pathway since it wasn’t affected by the addition of a neutralizing IFNAR antibody or the complete lack of IFNAR expression. Furthermore, the same synergistic effect was also observed when a CXCL10 promoter reporter was examined. Although the responsive promoter region contains both ISRE and NFκB sites, western blot analysis indicated that the combined treatment of IFN γ and dsRNA significantly augmented NFκB but not STAT1 activation as compared to the single treatment. Therefore, we conclude that IFN γ and dsRNA act in concert to potentiate CXCL10 expression in airway epithelial cells via an NFκB-dependent but IFNAR-STAT independent pathway and it is at least partly regulated at the transcriptional level.


title: Excessive production and extreme editing of human metapneumovirus defective interfering RNA is associated with type I IFN induction

text: Type I IFN production is one of the hallmarks of host innate immune responses upon virus infection. Whilst most respiratory viruses carry IFN antagonists, reports on human metapneumovirus (HMPV) have been conflicting. Using deep sequencing, we have demonstrated that HMPV particles accumulate excessive amounts of defective interfering RNA (DIs) rapidly upon in vitro passage, and that these are associated with IFN induction. Importantly, the DIs were edited extensively; up to 70 % of the original A and T residues had mutated to G or C, respectively. Such high editing rates of viral RNA have not, to our knowledge, been reported before. Bioinformatics and PCR assays indicated that adenosine deaminase acting on RNA (ADAR) was the most likely editing enzyme. HMPV thus has an unusually high propensity to generate DIs, which are edited at an unprecedented high frequency. The conflicting published data on HMPV IFN induction and antagonism are probably explained by DIs in virus stocks. The interaction of HMPV DIs with the RNA-editing machinery and IFN responses warrants further investigation.


title: Choindroitinase ABC I-Mediated Enhancement of Oncolytic Virus Spread and Anti Tumor Efficacy: A Mathematical Model

text: Oncolytic viruses are genetically engineered viruses that are designed to kill cancer cells while doing minimal damage to normal healthy tissue. After being injected into a tumor, they infect cancer cells, multiply inside them, and when a cancer cell is killed they move on to spread and infect other cancer cells. Chondroitinase ABC (Chase-ABC) is a bacterial enzyme that can remove a major glioma ECM component, chondroitin sulfate glycosoamino glycans from proteoglycans without any deleterious effects in vivo. It has been shown that Chase-ABC treatment is able to promote the spread of the viruses, increasing the efficacy of the viral treatment. In this paper we develop a mathematical model to investigate the effect of the Chase-ABC on the treatment of glioma by oncolytic viruses (OV). We show that the model's predictions agree with experimental results for a spherical glioma. We then use the model to test various treatment options in the heterogeneous microenvironment of the brain. The model predicts that separate injections of OV, one into the center of the tumor and another outside the tumor will result in better outcome than if the total injection is outside the tumor. In particular, the injection of the ECM-degrading enzyme (Chase-ABC) on the periphery of the main tumor core need to be administered in an optimal strategy in order to infect and eradicate the infiltrating glioma cells outside the tumor core in addition to proliferative cells in the bulk of tumor core. The model also predicts that the size of tumor satellites and distance between the primary tumor and multifocal/satellite lesions may be an important factor for the efficacy of the viral therapy with Chase treatment.


title: Significant cross reactive antibodies to influenza virus in adults and children during a period of marked antigenic drift

text: BACKGROUND: Little is known about the development of cross-reactive antibodies following natural exposure to pathogens. Such knowledge is critical in the development of new universal influenza vaccines. METHODS: To study the possibility of the presence of cross-reactive antibodies to influenza viruses which underwent a major antigenic drift between the years 1999 and 2007 sera from samples of 80 children and 400 adults were selected at random from the Israeli national serum bank. The sera was obtained in 2002 and in 2007, two time points that followed a major drift in the infectious H3N2 influenza virus strain (A/Panama/2007/99 to A/Wisconsin/67/2005). RESULTS: In the summer of 2002, 13% of the children had Hemagglutination Inhibition (HI) antibody titers of at least 40 and these antibodies recognized both A/Panama/2007/99 and A/Wisconsin/67/2005, where the latter strain only began to circulate in Israel in 2006. In 2007, 29% of the children had HI antibody titers of at least 40 directed against both A/Wisconsin/67/2005 and A/Panama/2007/99, even though they had never been exposed to the latter virus. Anti-A/Panama/2007/99 antibodies were detected in 58% and 68% of the 2002 and 2007 adult samples, respectively, while 8% and 39% had antibodies against A/Wisconsin/67/2005, respectively. CONCLUSIONS: The presence of naturally occurring cross-reactive influenza virus antibodies in a significant percentage of children has important implications for the development of a universal influenza vaccine.