title: Endogenous Cell Repair of Chronic Demyelination

text: In multiple sclerosis lesions, remyelination typically fails with repeated or chronic demyelinating episodes and results in neurologic disability. Acute demyelination models in rodents typically exhibit robust spontaneous remyelination that prevents appropriate evaluation of strategies for improving conditions of insufficient remyelination. In the current study, we used a mouse model of chronic demyelination induced by continuous ingestion of 0.2% cuprizone for 12 weeks. This chronic process depleted the oligodendrocyte progenitor population and impaired oligodendrocyte regeneration. Remyelination remained limited after removal of cuprizone from the diet. Fibroblast growth factor 2 (FGF2) expression was persistently increased in the corpus callosum of chronically demyelinated mice as compared with nonlesioned mice. We used FGF2(−/−)mice to determine whether removal of endogenous FGF2 promoted remyelination of chronically demyelinated areas. Wild-type and FGF2(−/−)mice exhibited similar demyelination during chronic cuprizone treatment. Importantly, in contrast to wild-type mice, the FGF2(−/−)mice spontaneously remyelinated completely during the recovery period after chronic demyelination. Increased remyelination in FGF2(−/−)mice correlated with enhanced oligodendroglial regeneration. FGF2 genotype did not alter the density of oligodendrocyte progenitor cells or proliferating cells after chronic demyelination. These findings indicate that attenuating FGF2 created a sufficiently permissive lesion environment for endogenous cells to effectively remyelinate viable axons even after chronic demyelination.


title: Automated identification of multiple micro-organisms from resequencing DNA microarrays

text: There is an increasing recognition that detailed nucleic acid sequence information will be useful and even required in the diagnosis, treatment and surveillance of many significant pathogens. Because generating detailed information about pathogens leads to significantly larger amounts of data, it is necessary to develop automated analysis methods to reduce analysis time and to standardize identification criteria. This is especially important for multiple pathogen assays designed to reduce assay time and costs. In this paper, we present a successful algorithm for detecting pathogens and reporting the maximum level of detail possible using multi-pathogen resequencing microarrays. The algorithm filters the sequence of base calls from the microarray and finds entries in genetic databases that most closely match. Taxonomic databases are then used to relate these entries to each other so that the microorganism can be identified. Although developed using a resequencing microarray, the approach is applicable to any assay method that produces base call sequence information. The success and continued development of this approach means that a non-expert can now perform unassisted analysis of the results obtained from partial sequence data.


title: Molecular dynamics simulations of human [Formula: see text]: the role of modified bases in mRNA recognition

text: Accuracy in translation of the genetic code into proteins depends upon correct tRNA–mRNA recognition in the context of the ribosome. In human [Formula: see text] three modified bases are present in the anticodon stem–loop—2-methylthio-N6-threonylcarbamoyladenosine at position 37 (ms(2)t(6)A37), 5-methoxycarbonylmethyl-2-thiouridine at position 34 (mcm(5)s(2)U34) and pseudouridine (ψ) at position 39—two of which, ms(2)t(6)A37 and mcm(5)s(2)U34, are required to achieve wild-type binding activity of wild-type human [Formula: see text] [C. Yarian, M. Marszalek, E. Sochacka, A. Malkiewicz, R. Guenther, A. Miskiewicz and P. F. Agris (2000) Biochemistry, 39, 13390–13395]. Molecular dynamics simulations of nine tRNA anticodon stem–loops with different combinations of nonstandard bases were performed. The wild-type simulation exhibited a canonical anticodon stair-stepped conformation. The ms(2)t(6) modification at position 37 is required for maintenance of this structure and reduces solvent accessibility of U36. Ms(2)t(6)A37 generally hydrogen bonds across the loop and may prevent U36 from rotating into solution. A water molecule does coordinate to ψ39 most of the simulation time but weakly, as most of the residence lifetimes are <40 ps.


title: A novel endonuclease IV post-PCR genotyping system

text: Here we describe a novel endonuclease IV (Endo IV) based assay utilizing a substrate that mimics the abasic lesions that normally occur in double-stranded DNA. The three component substrate is characterized by single-stranded DNA target, an oligonucleotide probe, separated from a helper oligonucleotide by a one base gap. The oligonucleotide probe contains a non-fluorescent quencher at the 5′ end and fluorophore attached to the 3′ end through a special rigid linker. Fluorescence of the oligonucleotide probe is efficiently quenched by the interaction of terminal dye and quencher when not hybridized. Upon hybridization of the oligonucleotide probe and helper probe to their complementary target, the phosphodiester linkage between the rigid linker and the 3′ end of the probe is efficiently cleaved, generating a fluorescent signal. In this study, the use of the Endo IV assay as a post-PCR amplification detection system is demonstrated. High sensitivity and specificity are illustrated using single nucleotide polymorphism detection.


title: Pandemic influenza preparedness: an ethical framework to guide decision-making

text: BACKGROUND: Planning for the next pandemic influenza outbreak is underway in hospitals across the world. The global SARS experience has taught us that ethical frameworks to guide decision-making may help to reduce collateral damage and increase trust and solidarity within and between health care organisations. Good pandemic planning requires reflection on values because science alone cannot tell us how to prepare for a public health crisis. DISCUSSION: In this paper, we present an ethical framework for pandemic influenza planning. The ethical framework was developed with expertise from clinical, organisational and public health ethics and validated through a stakeholder engagement process. The ethical framework includes both substantive and procedural elements for ethical pandemic influenza planning. The incorporation of ethics into pandemic planning can be helped by senior hospital administrators sponsoring its use, by having stakeholders vet the framework, and by designing or identifying decision review processes. We discuss the merits and limits of an applied ethical framework for hospital decision-making, as well as the robustness of the framework. SUMMARY: The need for reflection on the ethical issues raised by the spectre of a pandemic influenza outbreak is great. Our efforts to address the normative aspects of pandemic planning in hospitals have generated interest from other hospitals and from the governmental sector. The framework will require re-evaluation and refinement and we hope that this paper will generate feedback on how to make it even more robust.


title: Open lung biopsy in early-stage acute respiratory distress syndrome

text: INTRODUCTION: Acute respiratory distress syndrome (ARDS) has heterogeneous etiologies, rapid progressive change and a high mortality rate. To improve the outcome of ARDS, accurate diagnosis is essential to the application of effective early treatment. The present study investigated the clinical effects and safety of open lung biopsy (OLB) in patients with early-stage ARDS of suspected non-infectious origin. METHODS: We undertook a retrospective study of 41 patients with early-stage ARDS (defined as one week or less after intubation) who underwent OLB in two medical intensive care units of a tertiary care hospital from 1999 to 2005. Data analyzed included baseline characteristics, complication rate, pathological diagnoses, treatment alterations, and hospital survival. RESULTS: The age of patients was 55 ± 17 years (mean ± SD). The average ratio of arterial partial pressure of oxygen (PaO(2)) to fraction of inspired oxygen (FiO(2)) was 116 ± 43 mmHg (mean ± SD) at biopsy. Seventeen patients (41%) were immunocompromised. Postoperative complications occurred in 20% of patients (8/41). All biopsies provided a pathological diagnosis with a diagnostic yield of 100%. Specific pathological diagnoses were made for 44% of patients (18/41). Biopsy findings led to an alteration of treatment modality in 73% of patients (30/41). The treatment alteration rate was higher in patients with nonspecific diagnoses than in patients with specific diagnoses (p = 0.0024). Overall mortality was 50% (21/41) and was not influenced by age, gender, pre-OLB oxygenation, complication rate, pathological results, and alteration of treatment. There was no surgery-related mortality. The survival rate for immunocompromised patients was better than that for immunocompetent patients (71% versus 33%; p = 0.0187) in this study. CONCLUSION: Our retrospective study suggests that OLB was a useful and acceptably safe diagnostic procedure in some selected patients with early-stage ARDS.


title: Association and Host Selectivity in Multi-Host Pathogens

text: The distribution of multi-host pathogens over their host range conditions their population dynamics and structure. Also, host co-infection by different pathogens may have important consequences for the evolution of hosts and pathogens, and host-pathogen co-evolution. Hence it is of interest to know if the distribution of pathogens over their host range is random, or if there are associations between hosts and pathogens, or between pathogens sharing a host. To analyse these issues we propose indices for the observed patterns of host infection by pathogens, and for the observed patterns of co-infection, and tests to analyse if these patterns conform to randomness or reflect associations. Applying these tests to the prevalence of five plant viruses on 21 wild plant species evidenced host-virus associations: most hosts and viruses were selective for viruses and hosts, respectively. Interestingly, the more host-selective viruses were the more prevalent ones, suggesting that host specialisation is a successful strategy for multi-host pathogens. Analyses also showed that viruses tended to associate positively in co-infected hosts. The developed indices and tests provide the tools to analyse how strong and common are these associations among different groups of pathogens, which will help to understand and model the population biology of multi-host pathogens.


title: The Effectiveness of Contact Tracing in Emerging Epidemics

text: BACKGROUND: Contact tracing plays an important role in the control of emerging infectious diseases, but little is known yet about its effectiveness. Here we deduce from a generic mathematical model how effectiveness of tracing relates to various aspects of time, such as the course of individual infectivity, the (variability in) time between infection and symptom-based detection, and delays in the tracing process. In addition, the possibility of iteratively tracing of yet asymptomatic infecteds is considered. With these insights we explain why contact tracing was and will be effective for control of smallpox and SARS, only partially effective for foot-and-mouth disease, and likely not effective for influenza. METHODS AND FINDINGS: We investigate contact tracing in a model of an emerging epidemic that is flexible enough to use for most infections. We consider isolation of symptomatic infecteds as the basic scenario, and express effectiveness as the proportion of contacts that need to be traced for a reproduction ratio smaller than 1. We obtain general results for special cases, which are interpreted with respect to the likely success of tracing for influenza, smallpox, SARS, and foot-and-mouth disease epidemics. CONCLUSIONS: We conclude that (1) there is no general predictive formula for the proportion to be traced as there is for the proportion to be vaccinated; (2) variability in time to detection is favourable for effective tracing; (3) tracing effectiveness need not be sensitive to the duration of the latent period and tracing delays; (4) iterative tracing primarily improves effectiveness when single-step tracing is on the brink of being effective.


title: The Waiting Time for Inter-Country Spread of Pandemic Influenza

text: BACKGROUND: The time delay between the start of an influenza pandemic and its subsequent initiation in other countries is highly relevant to preparedness planning. We quantify the distribution of this random time in terms of the separate components of this delay, and assess how the delay may be extended by non-pharmaceutical interventions. METHODS AND FINDINGS: The model constructed for this time delay accounts for: (i) epidemic growth in the source region, (ii) the delay until an infected individual from the source region seeks to travel to an at-risk country, (iii) the chance that infected travelers are detected by screening at exit and entry borders, (iv) the possibility of in-flight transmission, (v) the chance that an infected arrival might not initiate an epidemic, and (vi) the delay until infection in the at-risk country gathers momentum. Efforts that reduce the disease reproduction number in the source region below two and severe travel restrictions are most effective for delaying a local epidemic, and under favourable circumstances, could add several months to the delay. On the other hand, the model predicts that border screening for symptomatic infection, wearing a protective mask during travel, promoting early presentation of cases arising among arriving passengers and moderate reduction in travel volumes increase the delay only by a matter of days or weeks. Elevated in-flight transmission reduces the delay only minimally. CONCLUSIONS: The delay until an epidemic of pandemic strain influenza is imported into an at-risk country is largely determined by the course of the epidemic in the source region and the number of travelers attempting to enter the at-risk country, and is little affected by non-pharmaceutical interventions targeting these travelers. Short of preventing international travel altogether, eradicating a nascent pandemic in the source region appears to be the only reliable method of preventing country-to-country spread of a pandemic strain of influenza.


title: Neutralizing Antibody Fails to Impact the Course of Ebola Virus Infection in Monkeys

text: Prophylaxis with high doses of neutralizing antibody typically offers protection against challenge with viruses producing acute infections. In this study, we have investigated the ability of the neutralizing human monoclonal antibody, KZ52, to protect against Ebola virus in rhesus macaques. This antibody was previously shown to fully protect guinea pigs from infection. Four rhesus macaques were given 50 mg/kg of neutralizing human monoclonal antibody KZ52 intravenously 1 d before challenge with 1,000 plaque-forming units of Ebola virus, followed by a second dose of 50 mg/kg antibody 4 d after challenge. A control animal was exposed to virus in the absence of antibody treatment. Passive transfer of the neutralizing human monoclonal antibody not only failed to protect macaques against challenge with Ebola virus but also had a minimal effect on the explosive viral replication following infection. We show that the inability of antibody to impact infection was not due to neutralization escape. It appears that Ebola virus has a mechanism of infection propagation in vivo in macaques that is uniquely insensitive even to high concentrations of neutralizing antibody.


title: Public health preparedness in Alberta: a systems-level study

text: BACKGROUND: Recent international and national events have brought critical attention to the Canadian public health system and how prepared the system is to respond to various types of contemporary public health threats. This article describes the study design and methods being used to conduct a systems-level analysis of public health preparedness in the province of Alberta, Canada. The project is being funded under the Health Research Fund, Alberta Heritage Foundation for Medical Research. METHODS/DESIGN: We use an embedded, multiple-case study design, integrating qualitative and quantitative methods to measure empirically the degree of inter-organizational coordination existing among public health agencies in Alberta, Canada. We situate our measures of inter-organizational network ties within a systems-level framework to assess the relative influence of inter-organizational ties, individual organizational attributes, and institutional environmental features on public health preparedness. The relative contribution of each component is examined for two potential public health threats: pandemic influenza and West Nile virus. DISCUSSION: The organizational dimensions of public health preparedness depend on a complex mix of individual organizational characteristics, inter-agency relationships, and institutional environmental factors. Our study is designed to discriminate among these different system components and assess the independent influence of each on the other, as well as the overall level of public health preparedness in Alberta. While all agree that competent organizations and functioning networks are important components of public health preparedness, this study is one of the first to use formal network analysis to study the role of inter-agency networks in the development of prepared public health systems.


title: The immunoregulatory and allergy-associated cytokines in the aetiology of the otitis media with effusion.

text: Inflammation in the middle ear mucosa, which can be provoked by different primary factors such as bacterial and viral infection, local allergic reactions and reflux, is the crucial event in the pathogenesis of otitis media with effusion (OME). Unresolved acute inflammatory responses or defective immunoregulation of middle inflammation can promote chronic inflammatory processes and stimulate the chronic condition of OME. Cytokines are the central molecular regulators of middle ear inflammation and can switch the acute phase of inflammation in the chronic stage and induce molecular-pathological processes leading to the histopathological changes accompanying OME. In this review we present cytokines identified in otitis media, immunoregulatory [interleukin (IL)-2, IL-10, transforming growth factor-beta]) and allergy associated (IL-4, IL-5, granulocyte-macrophage colony-stimulating factor), as crucial molecular regulators, responsible for chronic inflammation in the middle ear and the chronic condition of OME.


title: Bench-to-bedside review: Critical illness-associated cognitive dysfunction – mechanisms, markers, and emerging therapeutics

text: Cognitive dysfunction is common in critically ill patients, not only during the acute illness but also long after its resolution. A large number of pathophysiologic mechanisms are thought to underlie critical illness-associated cognitive dysfunction, including neuro-transmitter abnormalities and occult diffuse brain injury. Markers that could be used to evaluate the influence of specific mechanisms in individual patients include serum anticholinergic activity, certain brain proteins, and tissue sodium concentration determination via high-resolution three-dimensional magnetic resonance imaging. Although recent therapeutic advances in this area are exciting, they are still too immature to influence patient care. Additional research is needed if we are to understand better the relative contributions of specific mechanisms to the development of critical illness-associated cognitive dysfunction and to determine whether these mechanisms might be amenable to treatment or prevention.


title: ProCAT: a data analysis approach for protein microarrays

text: Protein microarrays provide a versatile method for the analysis of many protein biochemical activities. Existing DNA microarray analytical methods do not translate to protein microarrays due to differences between the technologies. Here we report a new approach, ProCAT, which corrects for background bias and spatial artifacts, identifies significant signals, filters nonspecific spots, and normalizes the resulting signal to protein abundance. ProCAT provides a powerful and flexible new approach for analyzing many types of protein microarrays.


title: Positional clustering improves computational binding site detection and identifies novel cis-regulatory sites in mammalian GABA(A) receptor subunit genes

text: Understanding transcription factor (TF) mediated control of gene expression remains a major challenge at the interface of computational and experimental biology. Computational techniques predicting TF-binding site specificity are frequently unreliable. On the other hand, comprehensive experimental validation is difficult and time consuming. We introduce a simple strategy that dramatically improves robustness and accuracy of computational binding site prediction. First, we evaluate the rate of recurrence of computational TFBS predictions by commonly used sampling procedures. We find that the vast majority of results are biologically meaningless. However clustering results based on nucleotide position improves predictive power. Additionally, we find that positional clustering increases robustness to long or imperfectly selected input sequences. Positional clustering can also be used as a mechanism to integrate results from multiple sampling approaches for improvements in accuracy over each one alone. Finally, we predict and validate regulatory sequences partially responsible for transcriptional control of the mammalian type A γ-aminobutyric acid receptor (GABA(A)R) subunit genes. Positional clustering is useful for improving computational binding site predictions, with potential application to improving our understanding of mammalian gene expression. In particular, predicted regulatory mechanisms in the mammalian GABA(A)R subunit gene family may open new avenues of research towards understanding this pharmacologically important neurotransmitter receptor system.


title: The Transmissibility of Highly Pathogenic Avian Influenza in Commercial Poultry in Industrialised Countries

text: BACKGROUND: With the increased occurrence of outbreaks of H5N1 worldwide there is concern that the virus could enter commercial poultry farms with severe economic consequences. METHODOLOGY/PRINCIPAL FINDINGS: We analyse data from four recent outbreaks of highly pathogenic avian influenza (HPAI) in commercial poultry to estimate the farm-to-farm reproductive number for HPAI. The reproductive number is a key measure of the transmissibility of HPAI at the farm level because it can be used to evaluate the effectiveness of the control measures. In these outbreaks the mean farm-to-farm reproductive number prior to controls ranged from 1.1 to 2.4, with the maximum farm-based reproductive number in the range 2.2 to 3.2. Enhanced bio-security, movement restrictions and prompt isolation of the infected farms in all four outbreaks substantially reduced the reproductive number, but it remained close to the threshold value 1 necessary to ensure the disease will be eradicated. CONCLUSIONS/SIGNIFICANCE: Our results show that depending on the particular situation in which an outbreak of avian influenza occurs, current controls might not be enough to eradicate the disease, and therefore a close monitoring of the outbreak is required. The method we used for estimating the reproductive number is straightforward to implement and can be used in real-time. It therefore can be a useful tool to inform policy decisions.


title: The influenza pandemic preparedness planning tool InfluSim

text: BACKGROUND: Planning public health responses against pandemic influenza relies on predictive models by which the impact of different intervention strategies can be evaluated. Research has to date rather focused on producing predictions for certain localities or under specific conditions, than on designing a publicly available planning tool which can be applied by public health administrations. Here, we provide such a tool which is reproducible by an explicitly formulated structure and designed to operate with an optimal combination of the competing requirements of precision, realism and generality. RESULTS: InfluSim is a deterministic compartment model based on a system of over 1,000 differential equations which extend the classic SEIR model by clinical and demographic parameters relevant for pandemic preparedness planning. It allows for producing time courses and cumulative numbers of influenza cases, outpatient visits, applied antiviral treatment doses, hospitalizations, deaths and work days lost due to sickness, all of which may be associated with economic aspects. The software is programmed in Java, operates platform independent and can be executed on regular desktop computers. CONCLUSION: InfluSim is an online available software which efficiently assists public health planners in designing optimal interventions against pandemic influenza. It can reproduce the infection dynamics of pandemic influenza like complex computer simulations while offering at the same time reproducibility, higher computational performance and better operability.


title: Immune pathways and defence mechanisms in honey bees Apis mellifera

text: Social insects are able to mount both group-level and individual defences against pathogens. Here we focus on individual defences, by presenting a genome-wide analysis of immunity in a social insect, the honey bee Apis mellifera. We present honey bee models for each of four signalling pathways associated with immunity, identifying plausible orthologues for nearly all predicted pathway members. When compared to the sequenced Drosophila and Anopheles genomes, honey bees possess roughly one-third as many genes in 17 gene families implicated in insect immunity. We suggest that an implied reduction in immune flexibility in bees reflects either the strength of social barriers to disease, or a tendency for bees to be attacked by a limited set of highly coevolved pathogens.


title: ElaD, a Deubiquitinating Protease Expressed by E. coli

text: BACKGROUND: Ubiquitin and ubiquitin-like proteins (Ubl) are designed to modify polypeptides in eukaryotes. Covalent binding of ubiquitin or Ubls to substrate proteins can be reversed by specific hydrolases. One particular set of cysteine proteases, the CE clan, which targets ubiquitin and Ubls, has homologs in eukaryotes, prokaryotes, and viruses. FINDINGS: We have cloned and analyzed the E. coli protein elaD, which is distantly related to eukaryotic CE clan members of the ULP/SENP protease family that are specific for SUMO and Nedd8. Previously misannotated as a putative sulfatase/phosphatase, elaD is an efficient and specific deubiquitinating enzyme in vitro. Interestingly, elaD is present in all intestinal pathogenic E. coli strains, but conspicuously absent from extraintestinal pathogenic strains (ExPECs). Further homologs of this protease can be found in Acanthamoeba Polyphaga Mimivirus, and in Alpha-, Beta-and Gammaproteobacteria. CONCLUSION: The expression of ULP/SENP-related hydrolases in bacteria therefore extends to plant pathogens and medically relevant strains of Escherichia coli, Legionella pneumophila, Rickettsiae, Chlamydiae, and Salmonellae, in which the elaD ortholog sseL has recently been identified as a virulence factor with deubiquitinating activity. As a counterpoint, our phylogenetic and functional examination reveals that ancient eukaryotic ULP/SENP proteases also have the potential of ubiquitin-specific hydrolysis, suggesting an early common origin of this peptidase clan.


title: Mutational analysis of human CEACAM1: the potential of receptor polymorphism in increasing host susceptibility to bacterial infection

text: A common overlapping site on the N-terminal IgV-like domain of human carcinoembryonic antigen (CEA)-related cell adhesion molecules (CEACAMs) is targeted by several important human respiratory pathogens. These include Neisseria meningitidis (Nm) and Haemophilus influenzae (Hi) that can cause disseminated or persistent localized infections. To define the precise structural features that determine the binding of distinct pathogens with CEACAMs, we have undertaken molecular modelling and mutation of the receptor molecules at previously implicated key target residues required for bacterial binding. These include Ser-32, Tyr-34, Val-39, Gln-44 and Gln-89, in addition to Ile-91, the primary docking site for the pathogens. Most, but not all, of these residues located adjacent to each other in a previous N-domain model of human CEACAM1, which was based on REI, CD2 and CD4. In the current studies, we have refined this model based on the mouse CEACAM1 crystal structure, and observe that all of the above residues form an exposed continuous binding region on the N-domain. Examination of the model also suggested that substitution of two of these residues 34 and 89 could affect the accessibility of Ile-91 for ligand binding. By introducing selected mutations at the positions 91, 34 and 89, we confirmed the primary importance of Ile-91 in all bacterial binding to CEACAM1 despite the inter- and intraspecies structural differences between the bacterial CEACAM-binding ligands. The studies further indicated that the efficiency of binding was significantly enhanced for specific strains by mutations such as Y34F and Q89N, which also altered the hierarchy of Nm versus Hi strain binding. These studies imply that distinct polymorphisms in human epithelial CEACAMs have the potential to decrease or increase the risk of infection by the receptor-targeting pathogens.


title: Immune reconstitution inflammatory syndrome (IRIS): review of common infectious manifestations and treatment options

text: The immune reconstitution inflammatory syndrome (IRIS) in HIV-infected patients initiating antiretroviral therapy (ART) results from restored immunity to specific infectious or non-infectious antigens. A paradoxical clinical worsening of a known condition or the appearance of a new condition after initiating therapy characterizes the syndrome. Potential mechanisms for the syndrome include a partial recovery of the immune system or exuberant host immunological responses to antigenic stimuli. The overall incidence of IRIS is unknown, but is dependent on the population studied and its underlying opportunistic infectious burden. The infectious pathogens most frequently implicated in the syndrome are mycobacteria, varicella zoster, herpesviruses, and cytomegalovirus (CMV). No single treatment option exists and depends on the underlying infectious agent and its clinical presentation. Prospective cohort studies addressing the optimal screening and treatment of opportunistic infections in patients eligible for ART are currently being conducted. These studies will provide evidence for the development of treatment guidelines in order to reduce the burden of IRIS. We review the available literature on the pathogenesis and epidemiology of IRIS, and present treatment options for the more common infectious manifestations of this diverse syndrome and for manifestations associated with a high morbidity.


title: Global Surveillance of Emerging Influenza Virus Genotypes by Mass Spectrometry

text: BACKGROUND: Effective influenza surveillance requires new methods capable of rapid and inexpensive genomic analysis of evolving viral species for pandemic preparedness, to understand the evolution of circulating viral species, and for vaccine strain selection. We have developed one such approach based on previously described broad-range reverse transcription PCR/electrospray ionization mass spectrometry (RT-PCR/ESI-MS) technology. METHODS AND PRINCIPAL FINDINGS: Analysis of base compositions of RT-PCR amplicons from influenza core gene segments (PB1, PB2, PA, M, NS, NP) are used to provide sub-species identification and infer influenza virus H and N subtypes. Using this approach, we detected and correctly identified 92 mammalian and avian influenza isolates, representing 30 different H and N types, including 29 avian H5N1 isolates. Further, direct analysis of 656 human clinical respiratory specimens collected over a seven-year period (1999–2006) showed correct identification of the viral species and subtypes with >97% sensitivity and specificity. Base composition derived clusters inferred from this analysis showed 100% concordance to previously established clades. Ongoing surveillance of samples from the recent influenza virus seasons (2005–2006) showed evidence for emergence and establishment of new genotypes of circulating H3N2 strains worldwide. Mixed viral quasispecies were found in approximately 1% of these recent samples providing a view into viral evolution. CONCLUSION/SIGNIFICANCE: Thus, rapid RT-PCR/ESI-MS analysis can be used to simultaneously identify all species of influenza viruses with clade-level resolution, identify mixed viral populations and monitor global spread and emergence of novel viral genotypes. This high-throughput method promises to become an integral component of influenza surveillance.


title: Transmission Parameters of the 2001 Foot and Mouth Epidemic in Great Britain

text: Despite intensive ongoing research, key aspects of the spatial-temporal evolution of the 2001 foot and mouth disease (FMD) epidemic in Great Britain (GB) remain unexplained. Here we develop a Markov Chain Monte Carlo (MCMC) method for estimating epidemiological parameters of the 2001 outbreak for a range of simple transmission models. We make the simplifying assumption that infectious farms were completely observed in 2001, equivalent to assuming that farms that were proactively culled but not diagnosed with FMD were not infectious, even if some were infected. We estimate how transmission parameters varied through time, highlighting the impact of the control measures on the progression of the epidemic. We demonstrate statistically significant evidence for assortative contact patterns between animals of the same species. Predictive risk maps of the transmission potential in different geographic areas of GB are presented for the fitted models.


title: Efficient replication of pneumonia virus of mice (PVM) in a mouse macrophage cell line

text: Pneumonia virus of mice (PVM; family Paramyxoviridae, subfamily Pneumovirinae) is a natural respiratory pathogen of rodent species and an important new model for the study of severe viral bronchiolitis and pneumonia. However, despite high virus titers typically detected in infected mouse lung tissue in vivo, cell lines used routinely for virus propagation in vitro are not highly susceptible to PVM infection. We have evaluated several rodent and primate cell lines for susceptibility to PVM infection, and detected highest virus titers from infection of the mouse monocyte-macrophage RAW 264.7 cell line. Additionally, virus replication in RAW 264.7 cells induces the synthesis and secretion of proinflammatory cytokines relevant to respiratory virus disease, including tumor necrosis factor-α (TNF-α), interferon-β (IFN-β), macrophage inflammatory proteins 1α and 1β (MIP-1α and MIP-1β) and the functional homolog of human IL-8, mouse macrophage inflammatory peptide-2 (MIP-2). Identification and characterization of a rodent cell line that supports the replication of PVM and induces the synthesis of disease-related proinflammatory mediators will facilitate studies of molecular mechanisms of viral pathogenesis that will complement and expand on findings from mouse model systems.


title: Designing and conducting tabletop exercises to assess public health preparedness for manmade and naturally occurring biological threats

text: BACKGROUND: Since 2001, state and local health departments in the United States (US) have accelerated efforts to prepare for high-impact public health emergencies. One component of these activities has been the development and conduct of exercise programs to assess capabilities, train staff and build relationships. This paper summarizes lessons learned from tabletop exercises about public health emergency preparedness and about the process of developing, conducting, and evaluating them. METHODS: We developed, conducted, and evaluated 31 tabletop exercises in partnership with state and local health departments throughout the US from 2003 to 2006. Participant self evaluations, after action reports, and tabletop exercise evaluation forms were used to identify aspects of the exercises themselves, as well as public health emergency responses that participants found more or less challenging, and to highlight lessons learned about tabletop exercise design. RESULTS: Designing the exercises involved substantial collaboration with representatives from participating health departments to assure that the scenarios were credible, focused attention on local preparedness needs and priorities, and were logistically feasible to implement. During execution of the exercises, nearly all health departments struggled with a common set of challenges relating to disease surveillance, epidemiologic investigations, communications, command and control, and health care surge capacity. In contrast, performance strengths were more varied across participating sites, reflecting specific attributes of individual health departments or communities, experience with actual public health emergencies, or the emphasis of prior preparedness efforts. CONCLUSION: The design, conduct, and evaluation of the tabletop exercises described in this report benefited from collaborative planning that involved stakeholders from participating health departments and exercise developers and facilitators from outside the participating agencies. While these exercises identified both strengths and vulnerabilities in emergency preparedness, additional work is needed to develop reliable metrics to gauge exercise performance, inform follow-up action steps, and to develop re-evaluation exercise designs that assess the impact of post-exercise interventions.


title: Transcript-level annotation of Affymetrix probesets improves the interpretation of gene expression data

text: BACKGROUND: The wide use of Affymetrix microarray in broadened fields of biological research has made the probeset annotation an important issue. Standard Affymetrix probeset annotation is at gene level, i.e. a probeset is precisely linked to a gene, and probeset intensity is interpreted as gene expression. The increased knowledge that one gene may have multiple transcript variants clearly brings up the necessity of updating this gene-level annotation to a refined transcript-level. RESULTS: Through performing rigorous alignments of the Affymetrix probe sequences against a comprehensive pool of currently available transcript sequences, and further linking the probesets to the International Protein Index, we generated transcript-level or protein-level annotation tables for two popular Affymetrix expression arrays, Mouse Genome 430A 2.0 Array and Human Genome U133A Array. Application of our new annotations in re-examining existing expression data sets shows increased expression consistency among synonymous probesets and strengthened expression correlation between interacting proteins. CONCLUSION: By refining the standard Affymetrix annotation of microarray probesets from the gene level to the transcript level and protein level, one can achieve a more reliable interpretation of their experimental data, which may lead to discovery of more profound regulatory mechanism.


title: Selective redox regulation of cytokine receptor signaling by extracellular thioredoxin-1

text: The thiol-disulfide oxidoreductase thioredoxin-1 (Trx1) is known to be secreted by leukocytes and to exhibit cytokine-like properties. Extracellular effects of Trx1 require a functional active site, suggesting a redox-based mechanism of action. However, specific cell surface proteins and pathways coupling extracellular Trx1 redox activity to cellular responses have not been identified so far. Using a mechanism-based kinetic trapping technique to identify disulfide exchange interactions on the intact surface of living lymphocytes, we found that Trx1 catalytically interacts with a single principal target protein. This target protein was identified as the tumor necrosis factor receptor superfamily member 8 (TNFRSF8/CD30). We demonstrate that the redox interaction is highly specific for both Trx1 and CD30 and that the redox state of CD30 determines its ability to engage the cognate ligand and transduce signals. Furthermore, we confirm that Trx1 affects CD30-dependent changes in lymphocyte effector function. Thus, we conclude that receptor–ligand signaling interactions can be selectively regulated by an extracellular redox catalyst.


title: Screen for ISG15-crossreactive Deubiquitinases

text: BACKGROUND: The family of ubiquitin-like molecules (UbLs) comprises several members, each of which has sequence, structural, or functional similarity to ubiquitin. ISG15 is a homolog of ubiquitin in vertebrates and is strongly upregulated following induction by type I interferon. ISG15 can be covalently attached to proteins, analogous to ubiquitination and with actual support of ubiquitin conjugating factors. Specific proteases are able to reverse modification with ubiquitin or UbLs by hydrolyzing the covalent bond between their C-termini and substrate proteins. The tail regions of ubiquitin and ISG15 are identical and we therefore hypothesized that promiscuous deubiquitinating proteases (DUBs) might exist, capable of recognizing both ubiquitin and ISG15. RESULTS: We have cloned and expressed 22 human DUBs, representing the major clades of the USP protease family. Utilizing suicide inhibitors based on ubiquitin and ISG15, we have identified USP2, USP5 (IsoT1), USP13 (IsoT3), and USP14 as ISG15-reactive proteases, in addition to the bona fide ISG15-specific protease USP18 (UBP43). USP14 is a proteasome-associated DUB, and its ISG15 isopeptidase activity increases when complexed with the proteasome. CONCLUSIONS: By evolutionary standards, ISG15 is a newcomer among the UbLs and it apparently not only utilizes the conjugating but also the deconjugating machinery of its more established relative ubiquitin. Functional overlap between these two posttranslational modifiers might therefore be more extensive than previously appreciated and explain the rather innocuous phenotype of ISG15 null mice.


title: s-RT-MELT for rapid mutation scanning using enzymatic selection and real time DNA-melting: new potential for multiplex genetic analysis

text: The rapidly growing understanding of human genetic pathways, including those that mediate cancer biology and drug response, leads to an increasing need for extensive and reliable mutation screening on a population or on a single patient basis. Here we describe s-RT-MELT, a novel technology that enables highly expanded enzymatic mutation scanning in human samples for germline or low-level somatic mutations, or for SNP discovery. GC-clamp-containing PCR products from interrogated and wild-type samples are hybridized to generate mismatches at the positions of mutations over one or multiple sequences in-parallel. Mismatches are converted to double-strand breaks using a DNA endonuclease (Surveyor™) and oligonucleotide tails are enzymatically attached at the position of mutations. A novel application of PCR enables selective amplification of mutation-containing DNA fragments. Subsequently, melting curve analysis, on conventional or nano-technology real-time PCR platforms, detects the samples that contain mutations in a high-throughput and closed-tube manner. We apply s-RT-MELT in the screening of p53 and EGFR mutations in cell lines and clinical samples and demonstrate its advantages for rapid, multiplexed mutation scanning in cancer and for genetic variation screening in biology and medicine.


title: Rapid Identification of Malaria Vaccine Candidates Based on α-Helical Coiled Coil Protein Motif

text: To identify malaria antigens for vaccine development, we selected α-helical coiled coil domains of proteins predicted to be present in the parasite erythrocytic stage. The corresponding synthetic peptides are expected to mimic structurally “native” epitopes. Indeed the 95 chemically synthesized peptides were all specifically recognized by human immune sera, though at various prevalence. Peptide specific antibodies were obtained both by affinity-purification from malaria immune sera and by immunization of mice. These antibodies did not show significant cross reactions, i.e., they were specific for the original peptide, reacted with native parasite proteins in infected erythrocytes and several were active in inhibiting in vitro parasite growth. Circular dichroism studies indicated that the selected peptides assumed partial or high α-helical content. Thus, we demonstrate that the bioinformatics/chemical synthesis approach described here can lead to the rapid identification of molecules which target biologically active antibodies, thus identifying suitable vaccine candidates. This strategy can be, in principle, extended to vaccine discovery in a wide range of other pathogens.


title: FluGenome: a web tool for genotyping influenza A virus

text: Influenza A viruses are hosted by numerous avian and mammalian species, which have shaped their evolution into distinct lineages worldwide. The viral genome consists of eight RNA segments that are frequently exchanged between different viruses via a process known as genetic reassortment. A complete genotype nomenclature is essential to describe gene segment reassortment. Specialized bioinformatic tools to analyze reassortment are not available, which hampers progress in understanding its role in host range, virulence and transmissibility of influenza viruses. To meet this need, we have developed a nomenclature to name influenza A genotypes and implemented a web server, FluGenome (http://www.flugenome.org/), for the assignment of lineages and genotypes. FluGenome provides functions for the user to interrogate the database in different modalities and get detailed reports on lineages and genotypes. These features make FluGenome unique in its ability to automatically detect genotype differences attributable to reassortment events in influenza A virus evolution.


title: Influenza pandemic intervention planning using InfluSim: pharmaceutical and non- pharmaceutical interventions

text: BACKGROUND: Influenza pandemic preparedness plans are currently developed and refined on national and international levels. Much attention has been given to the administration of antiviral drugs, but contact reduction can also be an effective part of mitigation strategies and has the advantage to be not limited per se. The effectiveness of these interventions depends on various factors which must be explored by sensitivity analyses, based on mathematical models. METHODS: We use the freely available planning tool InfluSim to investigate how pharmaceutical and non-pharmaceutical interventions can mitigate an influenza pandemic. In particular, we examine how intervention schedules, restricted stockpiles and contact reduction (social distancing measures and isolation of cases) determine the course of a pandemic wave and the success of interventions. RESULTS: A timely application of antiviral drugs combined with a quick implementation of contact reduction measures is required to substantially protract the peak of the epidemic and reduce its height. Delays in the initiation of antiviral treatment (e.g. because of parsimonious use of a limited stockpile) result in much more pessimistic outcomes and can even lead to the paradoxical effect that the stockpile is depleted earlier compared to early distribution of antiviral drugs. CONCLUSION: Pharmaceutical and non-pharmaceutical measures should not be used exclusively. The protraction of the pandemic wave is essential to win time while waiting for vaccine development and production. However, it is the height of the peak of an epidemic which can easily overtax general practitioners, hospitals or even whole public health systems, causing bottlenecks in basic and emergency medical care.


title: DNA Vaccines against Protozoan Parasites: Advances and Challenges

text: Over the past 15 years, DNA vaccines have gone from a scientific curiosity to one of the most dynamic research field and may offer new alternatives for the control of parasitic diseases such as leishmaniasis and Chagas disease. We review here some of the advances and challenges for the development of DNA vaccines against these diseases. Many studies have validated the concept of using DNA vaccines for both protection and therapy against these protozoan parasites in a variety of mouse models. The challenge now is to translate what has been achieved in these models into veterinary or human vaccines of comparable efficacy. Also, genome-mining and new antigen discovery strategies may provide new tools for a more rational search of novel vaccine candidates.


title: Estimating Individual and Household Reproduction Numbers in an Emerging Epidemic

text: Reproduction numbers, defined as averages of the number of people infected by a typical case, play a central role in tracking infectious disease outbreaks. The aim of this paper is to develop methods for estimating reproduction numbers which are simple enough that they could be applied with limited data or in real time during an outbreak. I present a new estimator for the individual reproduction number, which describes the state of the epidemic at a point in time rather than tracking individuals over time, and discuss some potential benefits. Then, to capture more of the detail that micro-simulations have shown is important in outbreak dynamics, I analyse a model of transmission within and between households, and develop a method to estimate the household reproduction number, defined as the number of households infected by each infected household. This method is validated by numerical simulations of the spread of influenza and measles using historical data, and estimates are obtained for would-be emerging epidemics of these viruses. I argue that the household reproduction number is useful in assessing the impact of measures that target the household for isolation, quarantine, vaccination or prophylactic treatment, and measures such as social distancing and school or workplace closures which limit between-household transmission, all of which play a key role in current thinking on future infectious disease mitigation.


title: Natural Killer Cells Promote Early CD8 T Cell Responses against Cytomegalovirus

text: Understanding the mechanisms that help promote protective immune responses to pathogens is a major challenge in biomedical research and an important goal for the design of innovative therapeutic or vaccination strategies. While natural killer (NK) cells can directly contribute to the control of viral replication, whether, and how, they may help orchestrate global antiviral defense is largely unknown. To address this question, we took advantage of the well-defined molecular interactions involved in the recognition of mouse cytomegalovirus (MCMV) by NK cells. By using congenic or mutant mice and wild-type versus genetically engineered viruses, we examined the consequences on antiviral CD8 T cell responses of specific defects in the ability of the NK cells to control MCMV. This system allowed us to demonstrate, to our knowledge for the first time, that NK cells accelerate CD8 T cell responses against a viral infection in vivo. Moreover, we identify the underlying mechanism as the ability of NK cells to limit IFN-α/β production to levels not immunosuppressive to the host. This is achieved through the early control of cytomegalovirus, which dramatically reduces the activation of plasmacytoid dendritic cells (pDCs) for cytokine production, preserves the conventional dendritic cell (cDC) compartment, and accelerates antiviral CD8 T cell responses. Conversely, exogenous IFN-α administration in resistant animals ablates cDCs and delays CD8 T cell activation in the face of NK cell control of viral replication. Collectively, our data demonstrate that the ability of NK cells to respond very early to cytomegalovirus infection critically contributes to balance the intensity of other innate immune responses, which dampens early immunopathology and promotes optimal initiation of antiviral CD8 T cell responses. Thus, the extent to which NK cell responses benefit the host goes beyond their direct antiviral effects and extends to the prevention of innate cytokine shock and to the promotion of adaptive immunity.


title: The Trojan Chicken Study, Minnesota

text: We conducted a study in the summer of 2004 at county fairs in the Midwest to investigate the role poultry exhibits have in spreading avian pathogens to humans. A nearly invisible powder (pathogen surrogate) that fluoresces under UV light was surreptitiously sprinkled each day on 1 show bird at each of 2 fairs. A UV light box was used to daily examine the hands of 94 poultry-exhibit participants (blinded regarding UV box results) for up to 4 days during the poultry shows. Enrollment and end-of-study questionnaires collected data on pathogen risk factors. Eight (8.5%) of 94 participants had evidence of fluorescent powder contamination (95% confidence interval 2.76%–14.26%). This contamination and infrequent handwashing practices suggest that county fairs are a possible venue for animal-to-human pathogen transmission.


title: The Restriction of Zoonotic PERV Transmission by Human APOBEC3G

text: The human APOBEC3G protein is an innate anti-viral factor that can dominantly inhibit the replication of some endogenous and exogenous retroviruses. The prospects of purposefully harnessing such an anti-viral defense are under investigation. Here, long-term co-culture experiments were used to show that porcine endogenous retrovirus (PERV) transmission from pig to human cells is reduced to nearly undetectable levels by expressing human APOBEC3G in virus-producing pig kidney cells. Inhibition occurred by a deamination-independent mechanism, likely after particle production but before the virus could immortalize by integration into human genomic DNA. PERV inhibition did not require the DNA cytosine deaminase activity of APOBEC3G and, correspondingly, APOBEC3G-attributable hypermutations were not detected. In contrast, over-expression of the sole endogenous APOBEC3 protein of pigs failed to interfere significantly with PERV transmission. Together, these data constitute the first proof-of-principle demonstration that APOBEC3 proteins can be used to fortify the innate anti-viral defenses of cells to prevent the zoonotic transmission of an endogenous retrovirus. These studies suggest that human APOBEC3G-transgenic pigs will provide safer, PERV-less xenotransplantation resources and that analogous cross-species APOBEC3-dependent restriction strategies may be useful for thwarting other endogenous as well as exogenous retrovirus infections.


title: Experimental infection of H5N1 HPAI in BALB/c mice

text: BACKGROUND: In 2005 huge epizooty of H5N1 HPAI occurred in Russia. It had been clear that territory of Russia becoming endemic for H5N1 HPAI. In 2006 several outbreaks have occurred. To develop new vaccines and antiviral therapies, animal models had to be investigated. We choose highly pathogenic strain for these studies. RESULTS: A/duck/Tuva/01/06 belongs to Quinghai-like group viruses. Molecular markers – cleavage site, K627 in PB2 characterize this virus as highly pathogenic. This data was confirmed by direct pathogenic tests: IVPI = 3.0, MLD(50 )= 1,4Log10EID(50). Also molecular analysis showed sensivity of the virus to adamantanes and neuraminidase inhibitors. Serological analysis showed wide cross-reactivity of this virus with sera produced to H5N1 HPAI viruses isolated earlier in South-East Asia. Mean time to death of infected animals was 8,19+/-0,18 days. First time acute delayed hemorrhagic syndrome was observed in mice lethal model. Hypercytokinemia was determined by elevated sera levels of IFN-gamma, IL-6, IL-10. CONCLUSION: Assuming all obtained data we can conclude that basic model parameters were characterized and virus A/duck/Tuva/01/06 can be used to evaluate anti-influenza vaccines and therapeutics.


title: Prediction of RNA Pseudoknots Using Heuristic Modeling with Mapping and Sequential Folding

text: Predicting RNA secondary structure is often the first step to determining the structure of RNA. Prediction approaches have historically avoided searching for pseudoknots because of the extreme combinatorial and time complexity of the problem. Yet neglecting pseudoknots limits the utility of such approaches. Here, an algorithm utilizing structure mapping and thermodynamics is introduced for RNA pseudoknot prediction that finds the minimum free energy and identifies information about the flexibility of the RNA. The heuristic approach takes advantage of the 5′ to 3′ folding direction of many biological RNA molecules and is consistent with the hierarchical folding hypothesis and the contact order model. Mapping methods are used to build and analyze the folded structure for pseudoknots and to add important 3D structural considerations. The program can predict some well known pseudoknot structures correctly. The results of this study suggest that many functional RNA sequences are optimized for proper folding. They also suggest directions we can proceed in the future to achieve even better results.


title: Cell-penetrating peptides as transporters for morpholino oligomers: effects of amino acid composition on intracellular delivery and cytotoxicity

text: Arginine-rich cell-penetrating peptides (CPPs) are promising transporters for intracellular delivery of antisense morpholino oligomers (PMO). Here, we determined the effect of L-arginine, D-arginine and non-α amino acids on cellular uptake, splice-correction activity, cellular toxicity and serum binding for 24 CPP−PMOs. Insertion of 6-aminohexanoic acid (X) or β-alanine (B) residues into oligoarginine R(8) decreased the cellular uptake but increased the splice-correction activity of the resulting compound, with a greater increase for the sequences containing more X residues. Cellular toxicity was not observed for any of the conjugates up to 10 μM. Up to 60 μM, only the conjugates with ⩾ 5 Xs exhibited time- and concentration-dependent toxicity. Substitution of L-arginine with D-arginine did not increase uptake or splice-correction activity. High concentration of serum significantly decreased the uptake and splice-correction activity of oligoarginine conjugates, but had much less effect on the conjugates containing X or B. In summary, incorporation of X/B into oligoarginine enhanced the antisense activity and serum-binding profile of CPP−PMO. Toxicity of X/B-containing conjugates was affected by the number of Xs, treatment time and concentration. More active, stable and less toxic CPPs can be designed by optimizing the position and number of R, D-R, X and B residues.


title: Lost in the World of Functional Genomics, Systems Biology, and Translational Research: Is There Life after the Milstein Award?

text: We've always wanted to save the world from the scourges of virus infection by developing better drugs and vaccines. But fully understanding the intricacies of virus-host interactions, the first step in achieving this goal, requires the ability to view the process on a grand scale. The advent of high-throughput technologies, such as DNA microarrays and mass spectrometry, provided the first opportunities to obtain such a view. Here we describe our efforts to use these tools to focus on the changes in cellular gene expression and protein abundance that occur in response to virus infection. By examining these changes in a comprehensive manner, we have been able to discover exciting new insights into innate immunity, interferon and cytokine signaling, and the strategies used by viruses to overcome these cellular defenses. Functional genomics may yet save the world from killer viruses.


title: Electrospray ionisation-cleavable tandem nucleic acid mass tag–peptide nucleic acid conjugates: synthesis and applications to quantitative genomic analysis using electrospray ionisation-MS/MS

text: The synthesis and characterization of isotopomer tandem nucleic acid mass tag–peptide nucleic acid (TNT–PNA) conjugates is described along with their use as electrospray ionisation-cleavable (ESI-Cleavable) hybridization probes for the detection and quantification of target DNA sequences by electrospray ionisation tandem mass spectrometry (ESI-MS/MS). ESI-cleavable peptide TNT isotopomers were introduced into PNA oligonucleotide sequences in a total synthesis approach. These conjugates were evaluated as hybridization probes for the detection and quantification of immobilized synthetic target DNAs using ESI-MS/MS. In these experiments, the PNA portion of the conjugate acts as a hybridization probe, whereas the peptide TNT is released in a collision-based process during the ionization of the probe conjugate in the electrospray ion source. The cleaved TNT acts as a uniquely resolvable marker to identify and quantify a unique target DNA sequence. The method should be applicable to a wide variety of assays requiring highly multiplexed, quantitative DNA/RNA analysis, including gene expression monitoring, genetic profiling and the detection of pathogens.


title: Hairpin structure within the 3′UTR of DNA polymerase β mRNA acts as a post-transcriptional regulatory element and interacts with Hax-1

text: Aberrant expression of DNA polymerase β, a key enzyme involved in base excision repair, leads to genetic instability and carcinogenesis. Pol β expression has been previously shown to be regulated at the level of transcription, but there is also evidence of post-transcriptional regulation, since rat transcripts undergo alternative polyadenylation, and the resulting 3′UTR contain at least one regulatory element. Data presented here indicate that RNA of the short 3′UTR folds to form a strong secondary structure (hairpin). Its regulatory role was established utilizing a luciferase-based reporter system. Further studies led to the identification of a protein factor, which binds to this element—the anti-apoptotic, cytoskeleton-related protein Hax-1. The results of in vitro binding analysis indicate that the formation of the RNA–protein complex is significantly impaired by disruption of the hairpin motif. We demonstrate that Hax-1 binds to Pol β mRNA exclusively in the form of a dimer. Biochemical analysis revealed the presence of Hax-1 in mitochondria, but also in the nuclear matrix, which, along with its transcript-binding properties, suggests that Hax-1 plays a role in post-transcriptional regulation of expression of Pol β.


title: Global public goods and the global health agenda: problems, priorities and potential

text: The 'global public good' (GPG) concept has gained increasing attention, in health as well as development circles. However, it has suffered in finding currency as a general tool for global resource mobilisation, and is at risk of being attached to almost anything promoting development. This overstretches and devalues the validity and usefulness of the concept. This paper first defines GPGs and describes the policy challenge that they pose. Second, it identifies two key areas, health R&D and communicable disease control, in which the GPG concept is clearly relevant and considers the extent to which it has been applied. We point out that that, while there have been many new initiatives, it is not clear that additional resources from non-traditional sources have been forthcoming. Yet achieving this is, in effect, the entire purpose of applying the GPG concept in global health. Moreover, the proliferation of disease-specific programs associated with GPG reasoning has tended to promote vertical interventions at the expense of more general health sector strengthening. Third, we examine two major global health policy initiatives, the Global Fund against AIDS, Tuberculosis and Malaria (GFATM) and the bundling of long-standing international health goals in the form of Millennium Development Goals (MDG), asking how the GPG perspective has contributed to defining objectives and strategies. We conclude that both initiatives are best interpreted in the context of traditional development assistance and, one-world rhetoric aside, have little to do with the challenge posed by GPGs for health. The paper concludes by considering how the GPG concept can be more effectively used to promote global health.


title: Outcome of paediatric intensive care survivors

text: The development of paediatric intensive care has contributed to the improved survival of critically ill children. Physical and psychological sequelae and consequences for quality of life (QoL) in survivors might be significant, as has been determined in adult intensive care unit (ICU) survivors. Awareness of sequelae due to the original illness and its treatment may result in changes in treatment and support during and after the acute phase. To determine the current knowledge on physical and psychological sequelae and the quality of life in survivors of paediatric intensive care, we undertook a computerised comprehensive search of online databases for studies reporting sequelae in survivors of paediatric intensive care. Studies reporting sequelae in paediatric survivors of cardiothoracic surgery and trauma were excluded, as were studies reporting only mortality. All other studies reporting aspects of physical and psychological sequelae were analysed. Twenty-seven studies consisting of 3,444 survivors met the selection criteria. Distinct physical and psychological sequelae in patients have been determined and seemed to interfere with quality of life. Psychological sequelae in parents seem to be common. Small numbers, methodological limitations and quantitative and qualitative heterogeneity hamper the interpretation of data. We conclude that paediatric intensive care survivors and their parents have physical and psychological sequelae affecting quality of life. Further well-designed prospective studies evaluating sequelae of the original illness and its treatment are warranted.


title: An evaluation of Comparative Genome Sequencing (CGS) by comparing two previously-sequenced bacterial genomes

text: BACKGROUND: With the development of new technology, it has recently become practical to resequence the genome of a bacterium after experimental manipulation. It is critical though to know the accuracy of the technique used, and to establish confidence that all of the mutations were detected. RESULTS: In order to evaluate the accuracy of genome resequencing using the microarray-based Comparative Genome Sequencing service provided by Nimblegen Systems Inc., we resequenced the E. coli strain W3110 Kohara using MG1655 as a reference, both of which have been completely sequenced using traditional sequencing methods. CGS detected 7 of 8 small sequence differences, one large deletion, and 9 of 12 IS element insertions present in W3110, but did not detect a large chromosomal inversion. In addition, we confirmed that CGS also detected 2 SNPs, one deletion and 7 IS element insertions that are not present in the genome sequence, which we attribute to changes that occurred after the creation of the W3110 lambda clone library. The false positive rate for SNPs was one per 244 Kb of genome sequence. CONCLUSION: CGS is an effective way to detect multiple mutations present in one bacterium relative to another, and while highly cost-effective, is prone to certain errors. Mutations occurring in repeated sequences or in sequences with a high degree of secondary structure may go undetected. It is also critical to follow up on regions of interest in which SNPs were not called because they often indicate deletions or IS element insertions.


title: Antibody-Based HIV-1 Vaccines: Recent Developments and Future Directions: A summary report from a Global HIV Vaccine Enterprise Working Group

text: The authors discuss humoral immune responses to HIV and approaches to designing vaccines that induce viral neutralizing and other potentially protective antibodies.


title: Host Gene Expression Profiling of Dengue Virus Infection in Cell Lines and Patients

text: BACKGROUND: Despite the seriousness of dengue-related disease, with an estimated 50–100 million cases of dengue fever and 250,000–500,000 cases of dengue hemorrhagic fever/dengue shock syndrome each year, a clear understanding of dengue pathogenesis remains elusive. Because of the lack of a disease model in animals and the complex immune interaction in dengue infection, the study of host response and immunopathogenesis is difficult. The development of genomics technology, microarray and high throughput quantitative PCR have allowed researchers to study gene expression changes on a much broader scale. We therefore used this approach to investigate the host response in dengue virus-infected cell lines and in patients developing dengue fever. METHODOLOGY/PRINCIPAL FINDINGS: Using microarray and high throughput quantitative PCR method to monitor the host response to dengue viral replication in cell line infection models and in dengue patient blood samples, we identified differentially expressed genes along three major pathways; NF-κB initiated immune responses, type I interferon (IFN) and the ubiquitin proteasome pathway. Among the most highly upregulated genes were the chemokines IP-10 and I-TAC, both ligands of the CXCR3 receptor. Increased expression of IP-10 and I-TAC in the peripheral blood of ten patients at the early onset of fever was confirmed by ELISA. A highly upregulated gene in the IFN pathway, viperin, was overexpressed in A549 cells resulting in a significant reduction in viral replication. The upregulation of genes in the ubiquitin-proteasome pathway prompted the testing of proteasome inhibitors MG-132 and ALLN, both of which reduced viral replication. CONCLUSION/SIGNIFICANCE: Unbiased gene expression analysis has identified new host genes associated with dengue infection, which we have validated in functional studies. We showed that some parts of the host response can be used as potential biomarkers for the disease while others can be used to control dengue viral replication, thus representing viable targets for drug therapy.


title: Species-specific evolution of immune receptor tyrosine based activation motif-containing CEACAM1-related immune receptors in the dog

text: BACKGROUND: Although the impact of pathogens on the evolution of the mammalian immune system is still under debate, proteins, which both regulate immune responses and serve as cellular receptors for pathogens should be at the forefront of pathogen-driven host evolution. The CEA (carcinoembryonic antigen) gene family codes for such proteins and indeed shows tremendous species-specific variation between human and rodents. Since little is known about the CEA gene family in other lineages of placental mammals, we expected to gain new insights into the evolution of the rapidly diverging CEA family by analyzing the CEA family of the dog. RESULTS: Here we describe the complete CEA gene family in the dog. We found that the gene coding for the ITIM-bearing immunoregulatory molecule CEACAM1 gave rise to a recent expansion of the canine CEA gene family by gene duplication, similar to that previously found in humans and mice. However, while the murine and human CEACAMs (carcinoembryonic antigen-related cell adhesion molecules) are predominantly secreted and GPI-anchored, respectively, in the dog, most of the CEACAMs represent ITAM-bearing transmembrane proteins. One of these proteins, CEACAM28, exhibits nearly complete sequence identity with the ligand-binding N domain of CEACAM1, but antagonizing signaling motifs in the cytoplasmic tail. Comparison of nonsynonymous and synonymous substitutions indicates that the CEACAM28 N domain is under the strongest purifying selection of all canine CEACAM1-related CEACAMs. In addition, CEACAM28 shows a similar expression pattern in resting immune cells and tissues as CEACAM1. However, upon activation CEACAM28 mRNA and CEACAM1 mRNA are differentially regulated. CONCLUSION: Thus, CEACAM1 and CEACAM28 are the first paired immune receptors identified within the CEA gene family, which are expressed on T cells and are most likely involved in the fine-tuning of T cell responses. The direction of gene conversion accompanied by purifying selection and expression in immune cells suggests the possibility that CEACAM28 evolved in response to selective pressure imposed by species-specific pathogens.


title: Nucleolus: the fascinating nuclear body

text: Nucleoli are the prominent contrasted structures of the cell nucleus. In the nucleolus, ribosomal RNAs are synthesized, processed and assembled with ribosomal proteins. RNA polymerase I synthesizes the ribosomal RNAs and this activity is cell cycle regulated. The nucleolus reveals the functional organization of the nucleus in which the compartmentation of the different steps of ribosome biogenesis is observed whereas the nucleolar machineries are in permanent exchange with the nucleoplasm and other nuclear bodies. After mitosis, nucleolar assembly is a time and space regulated process controlled by the cell cycle. In addition, by generating a large volume in the nucleus with apparently no RNA polymerase II activity, the nucleolus creates a domain of retention/sequestration of molecules normally active outside the nucleolus. Viruses interact with the nucleolus and recruit nucleolar proteins to facilitate virus replication. The nucleolus is also a sensor of stress due to the redistribution of the ribosomal proteins in the nucleoplasm by nucleolus disruption. The nucleolus plays several crucial functions in the nucleus: in addition to its function as ribosome factory of the cells it is a multifunctional nuclear domain, and nucleolar activity is linked with several pathologies. Perspectives on the evolution of this research area are proposed.


title: Antidiabetes and Anti-obesity Activity of Lagerstroemia speciosa

text: The leaves of Lagerstroemia speciosa (Lythraceae), a Southeast Asian tree more commonly known as banaba, have been traditionally consumed in various forms by Philippinos for treatment of diabetes and kidney related diseases. In the 1990s, the popularity of this herbal medicine began to attract the attention of scientists worldwide. Since then, researchers have conducted numerous in vitro and in vivo studies that consistently confirmed the antidiabetic activity of banaba. Scientists have identified different components of banaba to be responsible for its activity. Using tumor cells as a cell model, corosolic acid was isolated from the methanol extract of banaba and shown to be an active compound. More recently, a different cell model and the focus on the water soluble fraction of the extract led to the discovery of other compounds. The ellagitannin Lagerstroemin was identified as an effective component of the banaba extract responsible for the activity. In a different approach, using 3T3-L1 adipocytes as a cell model and a glucose uptake assay as the functional screening method, Chen et al. showed that the banaba water extract exhibited an insulin-like glucose transport inducing activity. Coupling HPLC fractionation with a glucose uptake assay, gallotannins were identified in the banaba extract as components responsible for the activity, not corosolic acid. Penta-O-galloyl-glucopyranose (PGG) was identified as the most potent gallotannin. A comparison of published data with results obtained for PGG indicates that PGG has a significantly higher glucose transport stimulatory activity than Lagerstroemin. Chen et al. have also shown that PGG exhibits anti-adipogenic properties in addition to stimulating the glucose uptake in adipocytes. The combination of glucose uptake and anti-adipogenesis activity is not found in the current insulin mimetic drugs and may indicate a great therapeutic potential of PGG.


title: Transmissibility of the Influenza Virus in the 1918 Pandemic

text: BACKGROUND: With a heightened increase in concern for an influenza pandemic we sought to better understand the 1918 Influenza pandemic, the most devastating epidemic of the previous century. METHODOLOGY/PRINCIPAL FINDINGS: We use data from several communities in Maryland, USA as well as two ships that experienced well-documented outbreaks of influenza in 1918. Using a likelihood-based method and a nonparametric method, we estimate the serial interval and reproductive number throughout the course of each outbreak. This analysis shows the basic reproductive number to be slightly lower in the Maryland communities (between 1.34 and 3.21) than for the enclosed populations on the ships (R(0) = 4.97, SE = 3.31). Additionally the effective reproductive number declined to sub epidemic levels more quickly on the ships (within around 10 days) than in the communities (within 30–40 days). The mean serial interval for the ships was consistent (3.33, SE = 5.96 and 3.81, SE = 3.69), while the serial intervals in the communities varied substantially (between 2.83, SE = 0.53 and 8.28, SE = 951.95). CONCLUSIONS/SIGNIFICANCE: These results illustrate the importance of considering the population dynamics when making statements about the epidemiological parameters of Influenza. The methods that we employ for estimation of the reproductive numbers and the serial interval can be easily replicated in other populations and with other diseases.


title: Surfactant therapy for acute respiratory failure in children: a systematic review and meta-analysis

text: INTRODUCTION: Exogenous surfactant is used to treat acute respiratory failure in children, although the benefits and harms in this setting are not clear. The objective of the present systematic review is to assess the effect of exogenous pulmonary surfactant on all-cause mortality in children mechanically ventilated for acute respiratory failure. METHODS: We searched the MEDLINE, EMBASE, CINAHL and Ovid Healthstar databases, the bibliographies of included trials and review articles, conference proceedings and trial registries. We included prospective, randomized, controlled trials of pulmonary surfactant that enrolled intubated and mechanically ventilated children with acute respiratory failure. We excluded trials that exclusively enrolled neonates or patients with asthma. Two reviewers independently rated trials for inclusion, extracted data and assessed the methodologic quality. We quantitatively pooled the results of trials, where suitable, using a random effects model. RESULTS: Six trials randomizing 314 patients were included. Surfactant use reduced mortality (relative risk = 0.7, 95% confidence interval = 0.4 to 0.97, P = 0.04), was associated with increased ventilator-free days (weighted mean difference = 2.5 days, 95% confidence interval = 0.3 to 4.6 days, P = 0.02) and reduced the duration of ventilation (weighted mean difference = 2.3 days, 95% confidence interval = 0.1 to 4.4 days, P = 0.04). CONCLUSION: Surfactant use decreased mortality, was associated with more ventilator-free days and reduced the duration of ventilation. No serious adverse events were reported.


title: Clinical review: Update of avian influenza A infections in humans

text: Influenza A viruses have a wide host range for infection, from wild waterfowl to poultry to humans. Recently, the cross-species transmission of avian influenza A, particularly subtype H5N1, has highlighted the importance of the non-human subtypes and their incidence in the human population has increased over the past decade. During cross-species transmission, human disease can range from the asymptomatic to mild conjunctivitis to fulminant pneumonia and death. With these cases, however, the risk for genetic change and development of a novel virus increases, heightening the need for public health and hospital measures. This review discusses the epidemiology, host range, human disease, outcome, treatment, and prevention of cross-transmission of avian influenza A into humans.


title: Clinical review: Mass casualty triage – pandemic influenza and critical care

text: Worst case scenarios for pandemic influenza planning in the US involve over 700,000 patients requiring mechanical ventilation. UK planning predicts a 231% occupancy of current level 3 (intensive care unit) bed capacity. Critical care planners need to recognise that mortality is likely to be high and the risk to healthcare workers significant. Contingency planning should, therefore, be multi-faceted, involving a robust health command structure, the facility to expand critical care provision in terms of space, equipment and staff and cohorting of affected patients in the early stages. It should also be recognised that despite this expansion of critical care, demand will exceed supply and a process for triage needs to be developed that is valid, reproducible, transparent and consistent with distributive justice. We advocate the development and validation of physiological scores for use as a triage tool, coupled with candid public discussion of the process.


title: Avian influenza outbreak in Turkey through health personnel's views: a qualitative study

text: BACKGROUND: Avian influenza threatens public health worldwide because it is usually associated with severe illness and, consequently, a higher risk of death. During the first months of 2006, Turkey experienced its first human avian influenza epidemic. A total of 21 human cases were identified, 12 of which were confirmed by the National Institute for Medical Research. Nine of the cases, including the four fatal ones, were from the Dogubeyazit-Van region. This study aims to evaluate the efforts at the avian influenza outbreak control in the Van-Dogubeyazit region in 2006 through the experiences of health personnel. METHODS: We conducted in-depth interviews with seventeen key informants who took active roles during the avian influenza outbreak in East Turkey during the first months of 2006. We gathered information about the initial responses, the progress and management of the outbreak control, and the reactions of the health professionals and the public. The findings of the study are reported according to the topics that appeared through thematic analysis of the interview transcripts. RESULTS: Following the first suspected avian influenza cases, a Van Crisis Coordination Committee was formed as the coordinating and decision-making body and played an important role in the appropriate timing of decisions. The health and agriculture services could not be well coordinated owing to the lack of integrated planning in preparation for outbreak and of integrated surveillance programs. Traditional poultry practice together with the low socio-economic status of the people and the lack of health care access in the region seemed to be a major risk for animal to animal and animal to human transmission. The strengths and weaknesses of the present health system – primary health care services, national surveillance and notification systems, human resource and management – affected the inter organizational coordination during the outbreak. Open communication between the government and the public played an important part in overcoming difficulties. CONCLUSION: Although there were problems during the avian influenza outbreak in Turkey, the rapid responses of the central and regional health authorities and the performance of the health workers were the key points in controlling the epidemic. The lessons from this outbreak should provide an opportunity for integrating the preparation plans of the health and agricultural organizations, and for revising the surveillance system and enhancing the role of the primary health care services in controlling epidemic disease. Developing successful strategies based on knowledge and experience may play a valuable role in delaying an avian influenza pandemic.


title: Influenza activity in Europe during eight seasons (1999–2007): an evaluation of the indicators used to measure activity and an assessment of the timing, length and course of peak activity (spread) across Europe

text: BACKGROUND: The European Influenza Surveillance Scheme (EISS) has collected clinical and virological data on influenza since 1996 in an increasing number of countries. The EISS dataset was used to characterise important epidemiological features of influenza activity in Europe during eight winters (1999–2007). The following questions were addressed: 1) are the sentinel clinical reports a good measure of influenza activity? 2) how long is a typical influenza season in Europe? 3) is there a west-east and/or south-north course of peak activity ('spread') of influenza in Europe? METHODS: Influenza activity was measured by collecting data from sentinel general practitioners (GPs) and reports by national reference laboratories. The sentinel reports were first evaluated by comparing them to the laboratory reports and were then used to assess the timing and spread of influenza activity across Europe during eight seasons. RESULTS: We found a good match between the clinical sentinel data and laboratory reports of influenza collected by sentinel physicians (overall match of 72% for +/- 1 week difference). We also found a moderate to good match between the clinical sentinel data and laboratory reports of influenza from non-sentinel sources (overall match of 60% for +/- 1 week). There were no statistically significant differences between countries using ILI (influenza-like illness) or ARI (acute respiratory disease) as case definition. When looking at the peak-weeks of clinical activity, the average length of an influenza season in Europe was 15.6 weeks (median 15 weeks; range 12–19 weeks). Plotting the peak weeks of clinical influenza activity reported by sentinel GPs against the longitude or latitude of each country indicated that there was a west-east spread of peak activity (spread) of influenza across Europe in four winters (2001–2002, 2002–2003, 2003–2004 and 2004–2005) and a south-north spread in three winters (2001–2002, 2004–2005 and 2006–2007). CONCLUSION: We found that: 1) the clinical data reported by sentinel physicians is a valid indicator of influenza activity; 2) the length of influenza activity across the whole of Europe was surprisingly long, ranging from 12–19 weeks; 3) in 4 out of the 8 seasons, there was a west-east spread of influenza, in 3 seasons a south-north spread; not associated with type of dominant virus in those seasons.


title: Diagnosis and treatment of severe sepsis

text: The burden of infection in industrialized countries has prompted considerable effort to improve the outcomes of patients with sepsis. This has been formalized through the Surviving Sepsis Campaign 'bundles', derived from the recommendations of 11 professional societies, which have promoted global improvement in those practices whose primary goal it is to reduce sepsis-related death. However, difficulties remain in implementing all of the procedures recommended by the experts, despite the apparent pragmatism of those procedures. We summarize the main proposals made by the Surviving Sepsis Campaign and focus on the difficulties associated with making a proper diagnosis and supplying adequate treatment promptly to septic patients.


title: Simian virus 40 vectors for pulmonary gene therapy

text: BACKGROUND: Sepsis remains the leading cause of death in critically ill patients. One of the primary organs affected by sepsis is the lung, presenting as the Acute Respiratory Distress Syndrome (ARDS). Organ damage in sepsis involves an alteration in gene expression, making gene transfer a potential therapeutic modality. This work examines the feasibility of applying simian virus 40 (SV40) vectors for pulmonary gene therapy. METHODS: Sepsis-induced ARDS was established by cecal ligation double puncture (2CLP). SV40 vectors carrying the luciferase reporter gene (SV/luc) were administered intratracheally immediately after sepsis induction. Sham operated (SO) as well as 2CLP rats given intratracheal PBS or adenovirus expressing luciferase served as controls. Luc transduction was evaluated by in vivo light detection, immunoassay and luciferase mRNA detection by RT-PCR in tissue harvested from septic rats. Vector abundance and distribution into alveolar cells was evaluated using immunostaining for the SV40 VP1 capsid protein as well as by double staining for VP1 and for the surfactant protein C (proSP-C). Immunostaining for T-lymphocytes was used to evaluate the cellular immune response induced by the vector. RESULTS: Luc expression measured by in vivo light detection correlated with immunoassay from lung tissue harvested from the same rats. Moreover, our results showed vector presence in type II alveolar cells. The vector did not induce significant cellular immune response. CONCLUSION: In the present study we have demonstrated efficient uptake and expression of an SV40 vector in the lungs of animals with sepsis-induced ARDS. These vectors appear to be capable of in vivo transduction of alveolar type II cells and may thus become a future therapeutic tool.


title: Role of receptor polymorphism and glycosylation in syncytium induction and host range variation of ecotropic mouse gammaretroviruses

text: BACKGROUND: We previously identified unusual variants of Moloney and Friend ecotropic mouse gammaretroviruses that have altered host range and are cytopathic in cells of the wild mouse species Mus dunni. Cytopathicity was attributed to different amino acid substitutions at the same critical env residue involved in receptor interaction: S82F in the Moloney variant Spl574, and S84A in the Friend mouse leukemia virus F-S MLV. Because M. dunni cells carry a variant CAT-1 cell surface virus receptor (dCAT-1), we examined the role of this receptor variant in cytopathicity and host range. RESULTS: We expressed dCAT-1 or mCAT-1 of NIH 3T3 origin in cells that are not normally infectible with ecotropic MLVs and evaluated the transfectants for susceptibility to virus infection and to virus-induced syncytium formation. The dCAT-1 transfectants, but not the mCAT-1 transfectants, were susceptible to virus-induced cytopathicity, and this cytopathic response was accompanied by the accumulation of unintegrated viral DNA. The dCAT-1 transfectants, however, did not also reproduce the relative resistance of M. dunni cells to Moloney MLV, and the mCAT-1 transfectants did not show the relative resistance of NIH 3T3 cells to Spl574. Western analysis, use of glycosylation inhibitors and mutagenesis to remove receptor glycosylation sites identified a possible role for cell-specific glycosylation in the modulation of virus entry. CONCLUSION: Virus entry and virus-induced syncytium formation using the CAT-1 receptor are mediated by a small number of critical amino acid residues in receptor and virus Env. Virus entry is modulated by glycosylation of cellular proteins, and this effect is cell and virus-specific.


title: The presence of the TAR RNA structure alters the programmed -1 ribosomal frameshift efficiency of the human immunodeficiency virus type 1 (HIV-1) by modifying the rate of translation initiation

text: HIV-1 uses a programmed -1 ribosomal frameshift to synthesize the precursor of its enzymes, Gag-Pol. The frameshift efficiency that is critical for the virus replication, is controlled by an interaction between the ribosome and a specific structure on the viral mRNA, the frameshift stimulatory signal. The rate of cap-dependent translation initiation is known to be altered by the TAR RNA structure, present at the 5′ and 3′ end of all HIV-1 mRNAs. Depending upon its concentration, TAR activates or inhibits the double-stranded RNA-dependent protein kinase (PKR). We investigated here whether changes in translation initiation caused by TAR affect HIV-1 frameshift efficiency. CD4+ T cells and 293T cells were transfected with a dual-luciferase construct where the firefly luciferase expression depends upon the HIV-1 frameshift. Translation initiation was altered by adding TAR in cis or trans of the reporter mRNA. We show that HIV-1 frameshift efficiency correlates negatively with changes in the rate of translation initiation caused by TAR and mediated by PKR. A model is presented where changes in the rate of initiation affect the probability of frameshifting by altering the distance between elongating ribosomes on the mRNA, which influences the frequency of encounter between these ribosomes and the frameshift stimulatory signal.


title: Establishing a nationwide emergency department-based syndromic surveillance system for better public health responses in Taiwan

text: BACKGROUND: With international concern over emerging infectious diseases (EID) and bioterrorist attacks, public health is being required to have early outbreak detection systems. A disease surveillance team was organized to establish a hospital emergency department-based syndromic surveillance system (ED-SSS) capable of automatically transmitting patient data electronically from the hospitals responsible for emergency care throughout the country to the Centers for Disease Control in Taiwan (Taiwan-CDC) starting March, 2004. This report describes the challenges and steps involved in developing ED-SSS and the timely information it provides to improve in public health decision-making. METHODS: Between June 2003 and March 2004, after comparing various surveillance systems used around the world and consulting with ED physicians, pediatricians and internal medicine physicians involved in infectious disease control, the Syndromic Surveillance Research Team in Taiwan worked with the Real-time Outbreak and Disease Surveillance (RODS) Laboratory at the University of Pittsburgh to create Taiwan's ED-SSS. The system was evaluated by analyzing daily electronic ED data received in real-time from the 189 hospitals participating in this system between April 1, 2004 and March 31, 2005. RESULTS: Taiwan's ED-SSS identified winter and summer spikes in two syndrome groups: influenza-like illnesses and respiratory syndrome illnesses, while total numbers of ED visits were significantly higher on weekends, national holidays and the days of Chinese lunar new year than weekdays (p < 0.001). It also identified increases in the upper, lower, and total gastrointestinal (GI) syndrome groups starting in November 2004 and two clear spikes in enterovirus-like infections coinciding with the two school semesters. Using ED-SSS for surveillance of influenza-like illnesses and enteroviruses-related infections has improved Taiwan's pandemic flu preparedness and disease control capabilities. CONCLUSION: Taiwan's ED-SSS represents the first nationwide real-time syndromic surveillance system ever established in Asia. The experiences reported herein can encourage other countries to develop their own surveillance systems. The system can be adapted to other cultural and language environments for better global surveillance of infectious diseases and international collaboration.


title: Amino Acid Similarity Accounts for T Cell Cross-Reactivity and for “Holes” in the T Cell Repertoire

text: BACKGROUND: Cytotoxic T cell (CTL) cross-reactivity is believed to play a pivotal role in generating immune responses but the extent and mechanisms of CTL cross-reactivity remain largely unknown. Several studies suggest that CTL clones can recognize highly diverse peptides, some sharing no obvious sequence identity. The emerging realization in the field is that T cell receptors (TcR) recognize multiple distinct ligands. PRINCIPAL FINDINGS: First, we analyzed peptide scans of the HIV epitope SLFNTVATL (SFL9) and found that TCR specificity is position dependent and that biochemically similar amino acid substitutions do not drastically affect recognition. Inspired by this, we developed a general model of TCR peptide recognition using amino acid similarity matrices and found that such a model was able to predict the cross-reactivity of a diverse set of CTL epitopes. With this model, we were able to demonstrate that seemingly distinct T cell epitopes, i.e., ones with low sequence identity, are in fact more biochemically similar than expected. Additionally, an analysis of HIV immunogenicity data with our model showed that CTLs have the tendency to respond mostly to peptides that do not resemble self-antigens. CONCLUSIONS: T cell cross-reactivity can thus, to an extent greater than earlier appreciated, be explained by amino acid similarity. The results presented in this paper will help resolving some of the long-lasting discussions in the field of T cell cross-reactivity.


title: Distinguishing Molecular Features and Clinical Characteristics of a Putative New Rhinovirus Species, Human Rhinovirus C (HRV C)

text: BACKGROUND: Human rhinoviruses (HRVs) are the most frequently detected pathogens in acute respiratory tract infections (ARTIs) and yet little is known about the prevalence, recurrence, structure and clinical impact of individual members. During 2007, the complete coding sequences of six previously unknown and highly divergent HRV strains were reported. To catalogue the molecular and clinical features distinguishing the divergent HRV strains, we undertook, for the first time, in silico analyses of all available polyprotein sequences and performed retrospective reviews of the medical records of cases in which variants of the prototype strain, HRV-QPM, had been detected. METHODOLOGY/PRINCIPLE FINDINGS: Genomic analyses revealed that the six divergent strains, residing within a clade we previously called HRV A2, had the shortest polyprotein of all picornaviruses investigated. Structure-based amino acid alignments identified conserved motifs shared among members of the genus Rhinovirus as well as substantive deletions and insertions unique to the divergent strains. Deletions mostly affected regions encoding proteins traditionally involved in antigenicity and serving as HRV and HEV receptor footprints. Because the HRV A2 strains cannot yet be cultured, we created homology models of predicted HRV-QPM structural proteins. In silico comparisons confirmed that HRV-QPM was most closely related to the major group HRVs. HRV-QPM was most frequently detected in infants with expiratory wheezing or persistent cough who had been admitted to hospital and required supplemental oxygen. It was the only virus detected in 65% of positive individuals. These observations contributed to an objective clinical impact ranging from mild to severe. CONCLUSIONS: The divergent strains did not meet classification requirements for any existing species of the genus Rhinovirus or Enterovirus. HRV A2 strains should be partitioned into at least one new species, putatively called Human rhinovirus C, populated by members detected with high frequency, from individuals with respiratory symptoms requiring hospital admission.