Rapid inactivation of Ebola virus (EBOV) is crucial for high-throughput testing of clinical samples in low-. In response to this outbreak, the international community has deployed an increasing number of Ebola diagnostic laboratories into the main West African countries affected (Guinea, Liberia, and Sierra Leone). Rapid diagnosis of EVD in humans is critical in the management of this disease in outbreak situations, as it allows prompt isolation and the chance to provide the best supportive care to patients, which helps reduce the overall infection rate and break the transmission chain.The preferred clinical sample for testing for Ebola virus (EBOV), an enveloped negative-sense single-strand RNA virus, is EDTA-blood, serum, or plasma with the primary diagnostic technology being real-time PCR (2). Other sample types, such as swabs or urine, may also be received by a laboratory. EBOV is designated in the United Kingdom by the Advisory Committee on Dangerous Pathogens (ACDP) as a hazard group 4 pathogen that must be handled under containment level (CL) 4 standards (biosafety level 4 [BSL4] in other countries). As such stringent laboratory infrastructure and containment procedures are required to handle viable EBOV material, only a few laboratories in Europe and elsewhere are suitably equipped (3). Within the timelines and budgets available, it has been impractical to create this laboratory infrastructure in West Africa, and therefore diagnostic laboratories have relied on methods that rapidly inactivate EBOV prior to routine processing and testing of samples by PCR.Laboratory methods of EBOV inactivation include gamma irradiation (4), nanoemulsion (5), photoinducible alkylating agents (6), and UV radiation (7), but these methods are primarily used for research purposes and may not be practicable in an outbreak situation that is likely to involve a high number of samples but reduced capability for handling and manipulation. In this context, any inactivation method must also be compatible with the EBOV PCR diagnostic approach.The CDC recommends Triton X-100 and heat treatment for 1 h for diagnostic samples containing hemorrhagic fever viruses (8), and this method has been adopted by many laboratories for handling of samples that may contain EBOV (9). Heating (alone or with acetic acid) for 1 h at 60°C has also been shown to reduce the titer of EBOV (10). Other guidelines can be nonspecific, specifying only the need for inactivation but not suggesting how (11) or suggesting generic use of denaturing/lysis buffers and/or heat (12). In the United Kingdom, the Advisory Committee on Dangerous Pathogens guidelines state that samples from confirmed cases may be processed in a containment level 2 laboratory using routine autoanalyzers if a containment level 4 laboratory is not available and provided specific procedures are followed (13). Within these guidelines, which encompass the application of multiple clinical tests, there is no specific requirement to inactivate EBOV (or other viral hemorrhagic fever agents) within a sa...

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There are no widely available vaccines or antiviral drugs capable of protecting against infection with Venezuelan equine encephalitis virus (VEEV), although an adenovirus vector expressing VEEV structural proteins protects mice from challenge with VEEV and is potentially a vaccine suitable for human use. This work examines whether alpha interferon (IFN-α) could act as an adjuvant for the adenovirus-based vaccine. IFN-α was either expressed by a plasmid linked to the adenovirus vaccine or encoded by a separate adenovirus vector administered as a mixture with the vaccine. In contrast to previous reports with other vaccines, the presence of IFN-α reduced the antibody response to VEEV. When IFN-α was encoded by adenovirus, the lack of a VEEV-specific response was accompanied by an increase in the immune response to the adenovirus vector. IFN-α also plays a direct role in defence against virus infection, inducing the expression of a large number of antiviral proteins. Adenovirus-delivered IFN-α protected mice from VEEV disease when administered 24 h prior to challenge, but not when administered 6 h post-challenge, suggesting that up to 24 h is required for the development of the IFN-mediated antiviral response.

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BackgroundThere is currently a requirement for antiviral therapies capable of protecting against infection with Venezuelan equine encephalitis virus (VEEV), as a licensed vaccine is not available for general human use. Monoclonal antibodies are increasingly being developed as therapeutics and are potential treatments for VEEV as they have been shown to be protective in the mouse model of disease. However, to be truly effective, the antibody should recognise multiple strains of VEEV and broadly reactive monoclonal antibodies are rarely and only coincidentally isolated using classical hybridoma technology.ResultsIn this work, methods were developed to reliably derive broadly reactive murine antibodies. A phage library was created that expressed single chain variable fragments (scFv) isolated from mice immunised with multiple strains of VEEV. A broadly reactive scFv was identified and incorporated into a murine IgG2a framework. This novel antibody retained the broad reactivity exhibited by the scFv but did not possess virus neutralising activity. However, the antibody was still able to protect mice against VEEV disease induced by strain TrD when administered 24 h prior to challenge.ConclusionA monoclonal antibody possessing reactivity to a wide range of VEEV strains may be of benefit as a generic antiviral therapy. However, humanisation of the murine antibody will be required before it can be tested in humans.Crown Copyright © 2009

Molecular diagnostic strategies are gaining wider acceptance and use in dermatology and dermatopathology as more practitioners in this field develop an understanding of the principles and applications of genomic technologies. Molecular testing is facilitating more accurate diagnosis, staging, and prognostication, in addition to guiding the selection of appropriate treatment, monitoring of therapy, and identification of novel therapeutic targets, for a wide variety of skin diseases.

BackgroundEastern equine encephalitis virus is an alphavirus that naturally cycles between mosquitoes and birds or rodents in Eastern States of the US. Equine infection occurs by being bitten by cross-feeding mosquitoes, with a case fatality rate of up to 75% in humans during epizootic outbreaks. There are no licensed medical countermeasures, and with an anticipated increase in mortality when exposed by the aerosol route based on anecdotal human data and experimental animal data, it is important to understand the pathogenesis of this disease in pursuit of treatment options. This report details the clinical and pathological findings of mice infected with EEEV by the aerosol route, and use as a model for EEEV infection in humans.MethodsMice were exposed by the aerosol route to a dose range of EEEV to establish the median lethal dose. A pathogenesis study followed whereby mice were exposed to a defined dose of virus and sacrificed at time-points thereafter for histopathological analysis and virology.ResultsClinical signs of disease appeared within 2 days post challenge, culminating in severe clinical signs within 24 h, neuro-invasion and dose dependent lethality. EEEV was first detected in the lung 1 day post challenge, and by day 3 peak viral titres were observed in the brain, spleen and blood, corresponding with severe meningoencephalitis, indicative of encephalitic disease. Lethality follows severe neurological signs, and may be linked to a threshold level of virus replication in the brain. Effective medical countermeasures for EEEV may necessitate early inoculation to inhibit infection of the brain in zoonotic incidents, and be able to traverse the blood-brain barrier to sufficiently interrupt replication in the brain in cases of aerosol infection.ConclusionsThere is little human data on the hazard posed by aerosol infection with encephalitic alphaviruses, and use of EEEV as a bioweapon may be by the aerosol route. A well characterized model of aerosol exposure that recapitulates some of the most severe human clinical features is necessary to evaluate the efficacy of putative medical countermeasures, and to increase our understanding about how this route of infection induces such rapid neuro-invasion and resulting disease.