Introducing EbolaCheck: potential for point-of-need infectious disease diagnosis

The 2013–2015 Ebolavirus disease humanitarian crisis has spurred the development of laboratory-free, point-of-care nucleic acid testing solutions. EbolaCheck is an international consortium of public health, academic and biotechnology industry stakeholders aiming to deliver clinical molecular diagnostic standard-of-care testing suitable for the West African milieu within 12 months. In this article, the current status of the EbolaCheck platform is discussed in the context of the current regulatory framework. Presented here are future goals to achieve differential diagnosis of hemorrhagic fever disease from <5-μl of whole blood samples or mucosal biofluids, in a single tube process, under 40 min and with minimal operator training requirements.

The role of pneumococcal carbon metabolism in colonisation and invasive disease

Streptococcus pneumoniae is a common asymptomatic commensal of the human nasopharynx. However, it is better known as a threatening pathogen that causes serious diseases such as pneumonia, meningitis and sepsis, as well as other less severe but more prevalent infections (e.g. otitis media). With the increase of antibiotic resistance and the limited efficacy of vaccines, pneumococcal infections remain a major problem. Therefore, the discovery of new therapeutic targets and preventive drugs are in high demand.(...)

Coronary heart disease and cerebrovascular disease mortality in young adults: recent trends in Europe.

Background: Over the last two decades, mortality from coronary heart disease (CHD) and cerebrovascular disease (CVD) declined by about 30% in the European Union (EU). Design: We analyzed trends in CHD (X ICD codes: I20-I25) and CVD (X ICD codes: I60-I69) mortality in young adults (age 35-44 years) in the EU as a whole and in 12 selected European countries, over the period 1980-2007. Methods: Data were derived from the World Health Organization mortality database. With joinpoint regression analysis, we identified significant changes in trends and estimated average annual percent changes (AAPC). Results: CHD mortality rates at ages 35-44 years have decreased in both sexes since the 1980s for most countries, except for Russia (130/100,000 men and 24/100,000 women, in 2005-7). The lowest rates (around 9/100,000 men, 2/100,000 women) were in France, Italy and Sweden. In men, the steepest declines in mortality were in the Czech Republic (AAPC = -6.1%), the Netherlands (-5.2%), Poland (-4.5%), and England and Wales (-4.5%). Patterns were similar in women, though with appreciably lower rates. The AAPC in the EU was -3.3% for men (rate = 16.6/100,000 in 2005-7) and -2.1% for women (rate = 3.5/100,000). For CVD, Russian rates in 2005-7 were 40/100,000 men and 16/100,000 women, 5 to 10-fold higher than in most western European countries. The steepest declines were in the Czech Republic and Italy for men, in Sweden and the Czech Republic for women. The AAPC in the EU was -2.5% in both sexes, with steeper declines after the mid-late 1990s (rates = 6.4/100,000 men and 4.3/100,000 women in 2005-7). Conclusions: CHD and CVD mortality steadily declined in Europe, except in Russia, whose rates were 10 to 15-fold higher than those of France, Italy or Sweden. Hungary and Poland, and also Scotland, where CHD trends were less favourable than in other western European countries, also emerge as priorities for preventive interventions.

Genetic variants in novel pathways influence blood pressure and cardiovascular disease risk.

Blood pressure is a heritable trait influenced by several biological pathways and responsive to environmental stimuli. Over one billion people worldwide have hypertension (≥140 mm Hg systolic blood pressure or  ≥90 mm Hg diastolic blood pressure). Even small increments in blood pressure are associated with an increased risk of cardiovascular events. This genome-wide association study of systolic and diastolic blood pressure, which used a multi-stage design in 200,000 individuals of European descent, identified sixteen novel loci: six of these loci contain genes previously known or suspected to regulate blood pressure (GUCY1A3-GUCY1B3, NPR3-C5orf23, ADM, FURIN-FES, GOSR2, GNAS-EDN3); the other ten provide new clues to blood pressure physiology. A genetic risk score based on 29 genome-wide significant variants was associated with hypertension, left ventricular wall thickness, stroke and coronary artery disease, but not kidney disease or kidney function. We also observed associations with blood pressure in East Asian, South Asian and African ancestry individuals. Our findings provide new insights into the genetics and biology of blood pressure, and suggest potential novel therapeutic pathways for cardiovascular disease prevention.

LINGO1 and LINGO2 variants are associated with essential tremor and Parkinson disease.

Genetic variation in the leucine-rich repeat and Ig domain containing 1 gene (LINGO1) was recently associated with an increased risk of developing essential tremor (ET) and Parkinson disease (PD). Herein, we performed a comprehensive study of LINGO1 and its paralog LINGO2 in ET and PD by sequencing both genes in patients (ET, n=95; PD, n=96) and by examining haplotype-tagging single-nucleotide polymorphisms (tSNPs) in a multicenter North American series of patients (ET, n=1,247; PD, n= 633) and controls (n=642). The sequencing study identified six novel coding variants in LINGO1 (p.S4C, p.V107M, p.A277T, p.R423R, p.G537A, p.D610D) and three in LINGO2 (p.D135D, p.P217P, p.V565V), however segregation analysis did not support pathogenicity. The association study employed 16 tSNPs at the LINGO1 locus and 21 at the LINGO2 locus. One variant in LINGO1 (rs9652490) displayed evidence of an association with ET (odds ratio (OR) =0.63; P=0.026) and PD (OR=0.54; P=0.016). Additionally, four other tSNPs in LINGO1 and one in LINGO2 were associated with ET and one tSNP in LINGO2 associated with PD (P<0.05). Further analysis identified one tSNP in LINGO1 and two in LINGO2 which influenced age at onset of ET and two tSNPs in LINGO1 which altered age at onset of PD (P<0.05). Our results support a role for LINGO1 and LINGO2 in determining risk for and perhaps age at onset of ET and PD. Further studies are warranted to confirm these findings and to determine the pathogenic mechanisms involved.

All Hands on Deck: Transdisciplinary Approaches to Emerging Infectious Disease

The increasing burden of emerging infectious diseases worldwide confronts us with numerous challenges, including the imperative to design research and responses that are commensurate to understanding the complex social and ecological contexts in which infectious diseases occur. A diverse group of scientists met in Hawaii in March 2005 to discuss the linked social and ecological contexts in which infectious diseases emerge. A subset of the meeting was a group that focused on ‘‘transdisciplinary approaches’’ to integrating knowledge across and beyond academic disciplines in order to improve prevention and control of emerging infections. This article is based on the discussions of that group. Here, we outline the epidemiological legacy that has dominated infectious disease research and control up until now, and introduce the role of new, transdisciplinary and systems-based approaches to emerging infectious diseases.Wedescribe four cases of transboundary health issues and use them to discuss the potential benefits, as well as the inherent difficulties, in understanding the social–ecological contexts in which infectious diseases occur and of using transdisciplinary approaches to deal with them.

Use and abuse of mathematical models: an illustration from the 2001 foot and mouth disease epidemic in the United Kingdom

Foot and mouth disease (FMD) is a major threat, not only to countries whose economies rely on agricultural exports, but also to industrialised countries that maintain a healthy domestic livestock industry by eliminating major infectious diseases from their livestock populations. Traditional methods of controlling diseases such as FMD require the rapid detection and slaughter of infected animals, and any susceptible animals with which they may have been in contact, either directly or indirectly. During the 2001 epidemic of FMD in the United Kingdom (UK), this approach was supplemented by a culling policy driven by unvalidated predictive models. The epidemic and its control resulted in the death of approximately ten million animals, public disgust with the magnitude of the slaughter, and political resolve to adopt alternative options, notably including vaccination, to control any future epidemics. The UK experience provides a salutary warning of how models can be abused in the interests of scientific opportunism.

Factors required for the uridylylation of the foot-and-mouth disease virus 3B1, 3B2, and 3B3 peptides by the RNA-dependent RNA polymerase (3D(pol)) in vitro

The 5' terminus of picornavirus genomic RNA is covalently linked to the virus-encoded peptide 313 (VTg). Foot-and-mouth disease virus (FMDV) is unique in encoding and using 3 distinct forms of this peptide. These peptides each act as primers for RNA synthesis by the virus-encoded RNA polymerase 3D(pol). To act as the primer for positive-strand RNA synthesis, the 3B peptides have to be uridylylated to form VPgpU(pU). For certain picornaviruses, it has been shown that this reaction is achieved by the 3D(pol) in the presence of the 3CD precursor plus an internal RNA sequence termed a cis-acting replication element (cre). The FMDV ere has been identified previously to be within the 5' untranslated region, whereas all other picornavirus cre structures are within the viral coding region. The requirements for the in vitro uridylylation of each of the FMDV 313 peptides has now been determined, and the role of the FMDV ere (also known as the 3B-uridylylation site, or bus) in this reaction has been analyzed. The poly(A) tail does not act as a significant template for FMDV 3B uridylylation.

Infection of hybrid primula seeds by Botrytis cinerea and latent disease spread through the plants

Botrytis cinerea occurred commonly on cultivated Primula ×polyantha seed. The fungus was mostly on the outside of the seed but sometimes was present within the seed. The fungus frequently caused disease at maturity in plants grown from the seed, demonstrated by growing plants in a filtered airflow, isolated from other possible sources of infection. Young, commercially produced P. ×polyantha plants frequently had symptomless B. cinerea infections spread throughout the plants for up to 3 months, with symptoms appearing only at flowering. Single genetic individuals of B. cinerea, as determined by DNA fingerprinting, often were dispersed widely throughout an apparently healthy plant. Plants could, however, contain more than one isolate.

The gut microbiota in obesity and metabolic disease - a novel therapeutic target

Obesity is sweeping the westernized world at a rate which far outstrips human genomic evolution, highlighting the importance of the obesogenic environment. Diet is an important component of this obesogenic environment, with certain diets (high fat, high refined carbohydrates and sugar) predisposing to overweight. On the other hand, there are also foods shown to protect against obesity and the diseases of obesity, including whole plant foods, dairy products, dietary fibre and functional foods like probiotics, prebiotics and phytochemicals. Interestingly, many of these foods mediate their health-promoting activities through the gut microbiota. The human gut microbiota itself has recently been identified as a contributory factor in this obesogenic environment, with differences observed between lean and obese. Evidence from human studies indicates that important groups of fermentative bacteria differ in abundance between lean and obese. Recently it has been suggested that anomalous microbiota composition in infancy can predispose to overweight in later life, highlighting the important role of optimal microbiota successional development, and that – as observed in laboratory animals – the gut microbiota may contribute to the aetiology of obesity. In this review we will introduce the gut microbiota, describe its interactions with major dietary components and the host, and then go on to discuss evidence indicating that the gut microbiota may contribute to the obesogenic environment. Finally, we will explore possible strategies for modulating the composition and activity of the human gut microbiota which may impact on obesity or the metabolic diseases associated with obesity. (Nutritional Therapy & Metabolism 2009; 27: 113-33)

Efficient production of foot-and-mouth disease virus empty capsids in insect cells following down regulation of 3C protease activity

Foot-and-mouth disease virus (FMDV) is a significant economically and distributed globally pathogen of Artiodactyla. Current vaccines are chemically inactivated whole virus particles that require large-scale virus growth in strict bio-containment with the associated risks of accidental release or incomplete inactivation. Non-infectious empty capsids are structural mimics of authentic particles with no associated risk and constitute an alternate vaccine candidate. Capsids self-assemble from the processed virus structural proteins, VP0, VP3 and VP1, which are released from the structural protein precursor P1-2A by the action of the virus-encoded 3C protease. To date recombinant empty capsid assembly has been limited by poor expression levels, restricting the development of empty capsids as a viable vaccine. Here expression of the FMDV structural protein precursor P1-2A in insect cells is shown to be efficient but linkage of the cognate 3C protease to the C-terminus reduces expression significantly. Inactivation of the 3C enzyme in a P1-2A-3C cassette allows expression and intermediate levels of 3C activity resulted in efficient processing of the P1-2A precursor into the structural proteins which assembled into empty capsids. Expression was independent of the insect host cell background and leads to capsids that are recognised as authentic by a range of anti-FMDV bovine sera suggesting their feasibility as an alternate vaccine.

Syndromic approach to arboviral diagnostics for global travelers as a basis for infectious disease surveillance

Background Arboviruses have overlapping geographical distributions and can cause symptoms that coincide with more common infections. Therefore, arbovirus infections are often neglected by travel diagnostics. Here, we assessed the potential of syndrome-based approaches for diagnosis and surveillance of neglected arboviral diseases in returning travelers. Method To map the patients high at risk of missed clinical arboviral infections we compared the quantity of all arboviral diagnostic requests by physicians in the Netherlands, from 2009 through 2013, with a literature-based assessment of the travelers’ likely exposure to an arbovirus. Results 2153 patients, with travel and clinical history were evaluated. The diagnostic assay for dengue virus (DENV) was the most commonly requested (86%). Of travelers returning from Southeast Asia with symptoms compatible with chikungunya virus (CHIKV), only 55% were tested. For travelers in Europe, arbovirus diagnostics were rarely requested. Over all, diagnostics for most arboviruses were requested only on severe clinical presentation. Conclusion Travel destination and syndrome were used inconsistently for triage of diagnostics, likely resulting in vast under-diagnosis of arboviral infections of public health significance. This study shows the need for more awareness among physicians and standardization of syndromic diagnostic algorithms

Farmer attitudes and livestock disease: exploring citizenship behaviour and peer monitoring across two BVD control schemes in the UK

The eradication of BVD in the UK is technically possible but appears to be socially untenable. The following study explored farmer attitudes to BVD control schemes in relation to advice networks and information sharing, shared aims and goals, motivation and benefits of membership, notions of BVD as a priority disease and attitudes toward regulation. Two concepts from the organisational management literature framed the study: citizenship behaviour where actions of individuals support the collective good (but are not explicitly recognised as such) and peer to peer monitoring (where individuals evaluate other’s behaviour). Farmers from two BVD control schemes in the UK participated in the study: Orkney Livestock Association BVD Eradication Scheme and Norfolk and Suffolk Cattle Breeders Association BVD Eradication Scheme. In total 162 farmers participated in the research (109 in-scheme and 53 out of scheme). The findings revealed that group helping and information sharing among scheme members was low with a positive BVD status subject to social censure. Peer monitoring in the form of gossip with regard to the animal health status of other farms was high. Interestingly, farmers across both schemes supported greater regulation with regard to animal health, largely due to the mistrust of fellow farmers following voluntary disease control measures. While group cohesiveness varied across the two schemes, without continued financial inducements, longer-term sustainability is questionable

False-positive results of a rapid K39-based strip test and Chagas disease

Background: The definitive diagnosis of visceral. leishmaniasis (VL) requires invasive procedures with demonstration of amastigotes in tissue or promastigotes in culture. Unfortunately, these approaches require laboratory materials not available in poor countries where the disease is endemic. The correct diagnosis of VL is important, and made more difficult by the fact that several common tropical diseases such as malaria, disseminated tuberculosis, and enteric fever share the same clinical presentation. Serological tests have been developed to replace parasitological diagnosis in the field. A commercially available K39-based strip test for VL has been developed for this purpose. The endemic area of leishmaniasis in Brazil overlaps the endemic area of Chagas disease, a disease that can cause false-positive serological test results. The aim of this study was to evaluate the incidence of false-positive exams using a rapid test for VL in patients with Chagas disease. Methods: A rapid test based on the recombinant K39 antigen of Leishmania was used in: (1) 30 patients with confirmed Chagas disease, (2) 30 patients with a serological diagnosis of Chagas disease by ELISA, indirect immunofluorescence, indirect hemagglutination, and chemiluminescence, (3) 30 healthy patients from a non-endemic area as the control group, (4) 30 patients with confirmed VL, and (5) 20 patients with proved cutaneous leishmaniasis. Results: The sensitivity and specificity of the rapid strip test were 100% when compared with healthy volunteers and those with confirmed Chagas disease. One false-positive result occurred in the group with Chagas disease diagnosed by serological tests (specificity of 96%). Conclusion: The rapid test based on recombinant K39 is a useful diagnostic assay, and a false-positive result rarely occurs in patients with a serological diagnosis of Chagas disease. (C) 2008 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Transport of cryptotanshinone, a major active triterpenoid in Salvia miltiorrhiza Bunge widely used in the treatment of stroke and Alzheimers disease, across the blood-brain

Cryptotanshinone (CTS), a major constituent from the roots of Salvia miltiorrhiza (Danshen), is widely used in the treatment of coronary heart disease, stroke and less commonly Alzheimer's disease. Our recent study indicates that CTS is a substrate for Pglycoprotein (PgP/MDR1/ABCB1). This study has investigated the nature of the brain distribution of CTS across the brain-blood barrier (BBB) using several in vitro and in vivo rodent models. A polarized transport of CTS was found in rat primary microvascular endothelial cell (RBMVEC) monolayers, with facilitated efflux from the abluminal side to luminal side. Addition of a PgP (e.g. verapamil and quinidine) or multi-drug resistance protein 1/2 (MRP1/2) inhibitor (e.g. probenecid and MK-571) in both luminal and abluminal sides attenuated the polarized transport. In a bilateral in situ brain perfusion model, the uptake of CTS into the cerebrum increased from 0.52 ± 0.1% at 1 min to 11.13 ± 2.36 ml/100 g tissue at 30 min and was significantly greater than that of sucrose. Co-perfusion of a PgP/MDR1 (e.g. verapamil) or MRP1/2 inhibitor (e.g. probenecid) significantly increased the brain distribution of CTS by 35.1-163.6%. The brain levels of CTS were only about 21% of those in plasma, and were significantly increased when coadministered with verapamil or probenecid in rats. The brain levels of CTS in rats subjected to middle cerebral artery occlusion and rats treated with quinolinic acid (a neurotoxin) were about 2- to 2.5-fold higher than the control rats. Moreover, the brain levels in mdr1a(-/-) and mrp1(-/-) mice were 10.9- and 1.5-fold higher than those in the wild-type mice, respectively. Taken collectively, these findings indicate that PgP and Mrp1 limit the brain penetration of CTS in rodents, suggesting a possible role of PgP and MRP1 in limiting the brain penetration of CTS in patients and causing drug resistance to Danshen therapy and interactions with conventional drugs that are substrates of PgP and MRP1. Further studies are needed to explore the role of other drug transporters in restricting the brain penetration of CTS and the clinical relevance.