European Bat Lyssavirus Type 2 RNA in Myotis daubentonii

Organ distribution of European bat lyssavirus type 2 viral RNA in its reservoir host, Myotis daubentonii (Daubenton's bat), was measured with a novel quantitative reverse transcription–polymerase chain reaction assay. High levels of genomic RNA were found in the brain and were also detectable in the tongue, bladder, and stomach

Emerging Technologies for the Detection of Rabies Virus: Challenges and Hopes in the 21st Century

The diagnosis of rabies is routinely based on clinical and epidemiological information, especially when exposures are reported in rabies-endemic countries. Diagnostic tests using conventional assays that appear to be negative, even when undertaken late in the disease and despite the clinical diagnosis, have a tendency, at times, to be unreliable. These tests are rarely optimal and entirely dependent on the nature and quality of the sample supplied. In the course of the past three decades, the application of molecular biology has aided in the development of tests that result in a more rapid detection of rabies virus. These tests enable viral strain identification from clinical specimens. Currently, there are a number of molecular tests that can be used to complement conventional tests in rabies diagnosis. Indeed the challenges in the 21st century for the development of rabies diagnostics are not of a technical nature; these tests are available now. The challenges in the 21st century for diagnostic test developers are two-fold: firstly, to achieve internationally accepted validation of a test that will then lead to its acceptance by organisations globally. Secondly, the areas of the world where such tests are needed are mainly in developing regions where financial and logistical barriers prevent their implementation. Although developing countries with a poor healthcare infrastructure recognise that molecular-based diagnostic assays will be unaffordable for routine use, the cost/benefit ratio should still be measured. Adoption of rapid and affordable rabies diagnostic tests for use in developing countries highlights the importance of sharing and transferring technology through laboratory twinning between the developed and the developing countries. Importantly for developing countries, the benefit of molecular methods as tools is the capability for a differential diagnosis of human diseases that present with similar clinical symptoms. Antemortem testing for human rabies is now possible using molecular techniques. These barriers are not insurmountable and it is our expectation that if such tests are accepted and implemented where they are most needed, they will provide substantial improvements for rabies diagnosis and surveillance. The advent of molecular biology and new technological initiatives that combine advances in biology with other disciplines will support the development of techniques capable of high throughput testing with a low turnaround time for rabies diagnosis

Evolutionary History of Rabies in Ghana

Rabies virus (RABV) is enzootic throughout Africa, with the domestic dog (Canis familiaris) being the principal vector. Dog rabies is estimated to cause 24,000 human deaths per year in Africa, however, this estimate is still considered to be conservative. Two sub-Saharan African RABV lineages have been detected in West Africa. Lineage 2 is present throughout West Africa, whereas Africa 1a dominates in northern and eastern Africa, but has been detected in Nigeria and Gabon, and Africa 1b was previously absent from West Africa. We confirmed the presence of RABV in a cohort of 76 brain samples obtained from rabid animals in Ghana collected over an eighteen-month period (2007–2009). Phylogenetic analysis of the sequences obtained confirmed all viruses to be RABV, belonging to lineages previously detected in sub-Saharan Africa. However, unlike earlier reported studies that suggested a single lineage (Africa 2) circulates in West Africa, we identified viruses belonging to the Africa 2 lineage and both Africa 1 (a and b) sub-lineages. Phylogeographic Bayesian Markov chain Monte Carlo analysis of a 405 bp fragment of the RABV nucleoprotein gene from the 76 new sequences derived from Ghanaian animals suggest that within the Africa 2 lineage three clades co-circulate with their origins in other West African countries. Africa 1a is probably a western extension of a clade circulating in central Africa and the Africa 1b virus a probable recent introduction from eastern Africa. We also developed and tested a novel reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of RABV in African laboratories. This RT-LAMP was shown to detect both Africa 1 and 2 viruses, including its adaptation to a lateral flow device format for product visualization. These data suggest that RABV epidemiology is more complex than previously thought in West Africa and that there have been repeated introductions of RABV into Ghana. This analysis highlights the potential problems of individual developing nations implementing rabies control programmes in the absence of a regional programme

A universal real-time assay for the detection of Lyssaviruses

► Universal real-time PCR primer pair demonstrated to hybridize to and detect each of the known Lyssaviruses (including Rabies virus) with greater sensitivity than a standard pan-Lyssavirus hemi-nested RT-PCR typically used. ► Target sequences of bat derived virus species unavailable for analysis (Aravan-, Khujand-, Irkut-, West Caucasian bat- and Shimoni bat virus) were synthesized to produce oligonucleotides and the synthetic DNA was used as a target for primer hybridization

Conventional, gel-based PCR-assays for the detection of lyssavirus species other than RABV.

<p>Conventional, gel-based PCR-assays for the detection of lyssavirus species other than RABV.</p

Conventional, gel-based PCR-assays for the detection rabies virus.

<p>Conventional, gel-based PCR-assays for the detection rabies virus.</p

Real-time PCR-assays for the detection of lyssavirus species other than RABV.

<p>Real-time PCR-assays for the detection of lyssavirus species other than RABV.</p

Conventional, gel-based PCR-assays for the generic detection of all lyssavirus species.

<p>Conventional, gel-based PCR-assays for the generic detection of all lyssavirus species.</p