First international collaborative study to evaluate rabies antibody detection method for use in monitoring the effectiveness of oral vaccination programmes in fox and raccoon dog in Europe.

The most effective and sustainable method to control and eliminate rabies in wildlife is the oral rabies vaccination (ORV) of target species, namely foxes and raccoon dogs in Europe. According to WHO and OIE, the effectiveness of oral vaccination campaigns should be regularly assessed via disease surveillance and ORV antibody monitoring. Rabies antibodies are generally screened for in field animal cadavers, whose body fluids are often of poor quality. Therefore, the use of alternative methods such as the enzyme-linked immunosorbent assay (ELISA) has been proposed to improve reliability of serological results obtained on wildlife samples. We undertook an international collaborative study to determine if the commercial BioPro ELISA Rabies Ab kit is a reliable and reproducible tool for rabies serological testing. Our results reveal that the overall specificity evaluated on naive samples reached 96.7%, and the coefficients of concordance obtained for fox and raccoon dog samples were 97.2% and 97.5%, respectively. The overall agreement values obtained for the four marketed oral vaccines used in Europe were all equal to or greater than 95%. The coefficients of concordance obtained by laboratories ranged from 87.2% to 100%. The results of this collaborative study show good robustness and reproducibility of the BioPro ELISA Rabies Ab kit.

Comparative assay of fluorescent antibody test results among twelve European National Reference Laboratories using various anti-rabies conjugates.

Twelve National Reference Laboratories (NRLs) for rabies have undertaken a comparative assay to assess the comparison of fluorescent antibody test (FAT) results using five coded commercial anti-rabies conjugates (Biorad, Bioveta, Fujirebio, Millipore, and SIFIN conjugates). Homogenized positive brain tissues infected with various lyssavirus species as well as negative samples were analyzed blindly using a standardized FAT procedure. Conjugates B, C, D, and E were found to be significantly more effective than conjugate A for GS7 (French RABV) diluted samples (1/8 and 1/100) while the frequency of concordant results of conjugates C and D differ significantly from conjugates A, B and E for CVS 27. For detection of EBLV-1 strains, conjugates C and D also presented a significantly lower frequency of discordant results compared to conjugates A, B and E. Conjugates B, C and D were found to be significantly more effective than conjugates E and A for EBLV-2 and ABLV samples. In view of these results, conjugates C and D set themselves apart from the others and appeared as the most effective of this 5-panel conjugates. This study clearly demonstrates that the variability of conjugates used by National Reference Laboratories can potentially lead to discordant results and influence assay sensitivity. In case of false negative results this could have a dramatic impact if the animal under investigation is responsible for human exposure. To avoid such situations, confirmatory tests should be implemented.

Molecular epidemiology of rabies in Ukraine.

To investigate the circulation of rabies virus in Ukraine, 78 rabies virus isolates were acquired from 14 states in 2002 and 2008-2010 for characterization. Partial sequences of nucleoprotein (359 nt) and glycoprotein (344 nt) genes were compared with those from neighbouring countries. The analysis identified 39 unique nucleoprotein genes and two geographically distinct RV variants belonging to the cosmopolitan lineage. The Ukrainian samples were similar to the North-East European lineage (NEE) (n = 19) and Russian group C (n = 20). The group C viruses were mainly isolated in Eastern Ukraine, from 9 regions, and from two other regions in Western Ukraine, suggesting the presence of group C throughout the country. These group C viruses are intermixed in bordering regions along the Dnieper River with viruses of group NEE, which were mainly isolated in six regions in Western Ukraine. Both nucleoprotein and glycoprotein gene analyses suggested evidence for cross-border movements of rabies virus.

Rabies surveillance in Poland (1992-2006).

Data on the surveillance of rabies in Poland from 1992 to 2006 is presented. Between 1992 and 2006, 22,575 rabies cases were diagnosed. There were 3,963 (17.5%) cases recorded in domestic animals, 18,555 (82.2%) in wildlife and 57 (0.3%) in bats. The peak of the epizootic in wildlife was recorded in 1992 with 3,084 cases. There were 2,547 cases recorded in wildlife with 2,079 (81.6%) found in the red fox. In 1993, the decision was made to apply oral rabies vaccination (ORV) of the red fox as a method of rabies control. The programme was started in western part of Poland, which borders Germany, and extended eastward in the following years. ORV was undertaken twice peryear (spring and autumn). In 2002, the vaccine campaign covered the whole territory of Poland. After 13 years of ORV, rabies incidence decreased sharply, from 3,084 cases in 1992 to 82 in 2006 (a 97% decrease in the number of rabies cases). The epidemiological situation in 2006 points to the constant decrease of rabies cases as a result of ORV. The red fox remains the main reservoir of rabies. The progress made over the past decade demonstrates that Poland is meeting the requirements to eliminate rabies in terrestrial animals.

Molecular epidemiology of rabies viruses in Europe.

Several different strains of classical rabies virus co-circulate in Europe. In order to investigate the roles of the host species and topography on the molecular epidemiology of these viruses, a 400 bp region of the nucleoprotein gene was sequenced and compared with more than 500 European virus isolates. Viruses from 21 European countries were represented including some unique panels of archived isolates from the former Republic of Yugoslavia, Estonia, the Czech Republic, Poland and Austria. Phylogenetic analysis of 198 unique sequences demonstrated numerous groups of viruses clustered at both geographical and host-species levels.

Rabies surveillance, trends in animal rabies and human post-exposure treatment in Poland, 1990 -2004.

This paper describes recent changes in the epizootical and epidemiological situation of rabies in Poland. Analysis of routine surveillance data on animal cases and human post-exposure treatment was performed in order to examine the impact of introduction of cell culture vaccine for human use and the implementation of the fox immunisation programme. The success of the immunisation programme for wild animals has become evident during the past 3 years, as a 9-fold decrease in animal rabies cases has been observed. To date, however, the downward trend in animal rabies cases has had no effect on the frequency of administration of the post-exposure treatment for humans. Moreover, two cases of locally acquired human rabies have occurred in patients who did not receive post-exposure vaccination. These cases prove that rabies should be still considered a public health concern in Poland.