Bluetongue in ruminants, updates

Bluetongue is a vector-borne animal disease spread by midges of the Culicoides genus. The virus has a segmented genome structure of 10 double-stranded RNA molecules. The virus is also classified into serotypes of which more than 36 are known. The first 24 serotypes are notifiable according to the World organisation of Animal Health (WOAH).

Genetic analysis of the bluetongue virus (BTV) has shown that it was the same virus (BTV-3/NET2023) that was also detected a year earlier.This is the result of research by Wageningen Bioveterinary Research (WBVR, part of Wageningen University & Research) commissioned by the Dutch Ministry of Agriculture, Fishery, Food security and Nature (LVVN).

To determine the genetic composition, a so-called sequence analysis is done. The sequence is mapped at the level of RNA and virus proteins. All the sequences together form the complete genetic material of the virus. “Our research shows that the bluetongue virus found in the field in 2024 is practically identical to the virus we found in September/October 2023,” said WBVR researcher Marc Engelsma. The similarities are more than 99.9 percent at both nucleotide level (genetic material) and protein level (virus proteins).

For the immunogenic virus protein VP2, the similarity is even 100 per cent. “This VP2 protein is the most important immunogenic protein,” Engelsma explains. The body recognises such an immunogenic protein of a virus as foreign to the body, the immune system makes antibodies against that protein and thus tries to clear the virus. Vaccines are designed to stimulate or accelerate this process. The BTV-3 vaccines authorised for use in the Netherlands are based on killed-off BTV-3 that also contains this protein. “Based on this genetic analysis, we conclude that there are no changes in the virus itself that affect the efficacy of the three available vaccines authorised for use.”

The Netherlands first experienced bluetongue in 2006, serotype 8 at the time. Eventually, this serotype circulated until 2008, after which the outbreak died out following a voluntary vaccination campaign. Retrospectively, there also appear to have been some incidental introductions of serotypes 6 and 1 in 2008, with the last animal testing positive in 2009. The Netherlands was declared blue tongue-free in 2012.

Since September 2023, the Netherlands has again been affected by bluetongue, this time serotype 3.

All ruminants are susceptible to infection with the bluetongue virus, but the clinical signs vary by species. Sheep and cattle have severe signs of disease during infection with bluetongue. Goats, llamas, alpacas, mouflons, bisons, yaks and cervids can also become ill, but often less severely. Blue tongue virus was also detected in two dogs in 2023; these animals are considered final hosts of the virus.

At WBVR, genetic material of blue tongue virus can be detected by Polymerase Chain Reaction (PCR) and blue tongue-specific antibodies by enzyme-linked immunosorbent assay (ELISA). To demonstrate active blue tongue infection, a PCR is performed. An ELISA can be performed to investigate whether an animal has experienced blue tongue, or may have been vaccinated. Both PCR and ELISA are tests aimed at detecting the virus; these tests are not suitable for typing the virus and determining which serotype is active.

When a blood sample (EDTA blood tube) is sent in, it is first registered. The sample is then divided into several tubes to be processed in the laboratory. There, all nucleic acids (component of genetic material) present in the blood are isolated. These nucleic acids are then subjected to the specific PCR test for bluetongue. A positive result is confirmation that the genetic material of the blue tongue virus has been detected. This positive result (indicated by ‘CONF’ in a test result) is cause to do the whole analysis again, to ensure that the sample has gone through the correct procedure. Only if the sample's second PCR test is also positive will the submitted sample be given ‘demonstrated’ status.

How long does it take to get the result of an analysis?

WBVR aims to examine all samples as soon as possible. PCR analyses have an average turnaround time of 4 to 7 days.

Our bluetongue PCR is bluetongue-wide. PCR analysis only determines that genetic material of bluetongue virus is present in the sample, however, this analysis does not allow serotyping.

The letters and numbers BTV03 and BTV10 on the results report refer to product codes used, not to the serotyping of the virus. The product codes on the results form are added to show which test is involved. By way of explanation, BTV03 stands for ‘BTV-PCR’ and always has the result ‘-’ if the virus was not found, or ‘CONF’ if bluetongue virus was detected. The sample is then confirmed (= analysed for a second time); the product code for this is BTV10 (BTV PCR CONF). Here the result can be ‘Not demonstrated’ if the virus is absent or ‘Demonstrated’ if the virus has been detected again.

Unfortunately, it is not possible to change the product codes in the laboratory management system.