Peste des petits ruminants (PPR) is a contagious and economically important disease affecting production of small ruminants (i.e., sheep and goats). Taking into consideration the lessons learnt from the Global Rinderpest Eradication Programme (GREP), PPR is now targeted by the international veterinary community as the next animal disease to be eradicated. To support the African continental programme for the control of PPR, the Pan African Veterinary Vaccine Centre of the African Union (AU-PANVAC) is developing diagnostics tools. Here, we describe the development of a blocking enzyme-linked immunosorbent assay (bELISA) that allows testing of a large number of samples for specific detection of antibodies directed against PPR virus in sheep and goat sera. The PPR bELISA uses an anti-haemagglutinin (H) monoclonal antibody (MAb) as a competitor antibody, and tests results are interpreted using the percentage of inhibition (PI) of MAb binding generated by the serum sample. PI values below or equal to 18% (PI ≤ 18%) are negative, PI values greater than or equal to 25% (PI ≥ 25%) are positive, and PI values greater than 18% and below 25% are doubtful. The diagnostic specificity (DSp) and diagnostic sensitivity (DSe) were found to be 100% and 93.74%, respectively. The H-based PPR-bELISA showed good correlation with the virus neutralization test (VNT), the gold standard test, with a kappa value of 0.947. The H-based PPR-bELISA is more specific than the commercial kit ID Screen® PPR Competition (N-based PPR-cELISA) from IDvet (France), but the commercial kit is slightly more sensitive than the H-based PPR-bELISA. The validation process also indicated good repeatability and reproducibility of the H-based PPR-bELISA, making this new test a suitable tool for the surveillance and sero-monitoring of the vaccination campaign.
In December 2017, Peste des Petits Ruminants (PPR) emerged in Burundi (East Africa) and rapidly spread to five provinces (Gitega, Kirundo, Mwaro, Muramvya and Karuzi) in the country, causing severe disease and killing more than 4,000 goats in the province of Gitega alone. An initial outbreak investigation was conducted in December 2017 by the Burundi Government Veterinary Services and samples were collected for laboratory confirmation. A competitive Enzyme Linked Immuno‐Sorbent Assay (cELISA: Chinese Patent No. ZL201210278970.9) supplied by the Lanzhou Veterinary Research Institute was used to test 112 sera and results showed around 37.5% positive samples. This high level of PPR positive sera in an animal population where PPR infection and vaccination had not been previously reported indicated the exposure of the animals to PPRV. Subsequently in January 2018, the laboratory tests conducted at the African Union‐Pan African Veterinary Vaccine Centre (AU‐PANVAC) laboratories following a joint investigative mission by the African Union‐Interafrican Bureau for Animal Resources (AU‐IBAR), AU‐PANVAC and the East African Community (EAC) confirmed the presence of PPR in Burundi. Samples tested by conventional RT‐PCR indicated the presence of the PPR virus (PPRV). Confirmatory isolation of the virus was also performed. Phylogenetic analysis revealed that the virus belongs to lineage III and shows a close relationship with PPRV isolates from Kenya in 2011 and Uganda in 2012. A possible explanation for the outbreaks of PPR in Burundi between December 2017 and February 2018 is presented.
An effective contagious caprine pleuropneumonia (CCPP) vaccine is essential for the increased production of healthy goats in a cost-effective manner and the prevention of animal-to-animal transmission for both domestic animals and wildlife. Quality control of this vaccine ensures that a reliable supply of pure, safe, and potent batches is obtained. As part of this control, in vitro quantification of Mycoplasma capricolum subsp. capripneumoniae (Mccp) protein in the final vaccines is required before the CCPP vaccine undergoes batch release and certification. The current method used for quantification is based on the measurement of total protein using the bicinchonic acid (BCA) test. This method quantifies the total amount of protein in the vaccine including contaminant protein from media, which can lead to overestimation of the quantity of Mccp protein, resulting in reduced vaccine immunogenicity. An immuno-capture ELISA (ICE) was developed for specific detection and quantification of the Mccp antigen in the CCPP vaccine. As the ICE detects and measures the amount of antigen between two layers of antibodies, capture and detecting antibodies are required. Mouse monoclonal antibodies (mAbs) that detect the Mccp antigen were produced and characterized. One of these mAbs, Mccp-25, was used to develop the ICE as an unlabelled capture antibody and horseradish peroxidase conjugated detecting antibody. The ICE was standardized and evaluated using an internal reference sample, experimental CCPP vaccines and commercial CCPP vaccines. A comparison between the polymerase chain reaction (PCR) and ICE showed good correlation between the two assays. Also, an in vitro ICE method correlated well with an in vivo sero-conversion in goats that were vaccinated with selected test vaccines. The sensitivity of the ICE was estimated at 30 ng/ml.
Enzyme linked immunosorbent assays (ELISAs) have been developed for the detection of antibodies against contagious caprine pleuropneumonia (CCPP), the causative agent of which is Mycoplasma capricolum subsp. Capripneumoniae (Mccp). The currently available commercial CCPP competitive ELISA (CCPP cELISA) kit produced and supplied by IDEXX Company (Westbrook, Maine, United States) is relatively expensive for most African laboratories. To address this issue and provide a variety of choices, a sensitive and specific blocking-ELISA (b-ELISA) test to detect antibodies against CCPP was developed. We describe the newly developed CCPP blocking-ELISA based on the blocking of an epitope of a monoclonal antibody (Mccp-25) by a positive serum sample against the Mccp protein coated on a plate. The Percentage Inhibition (PI) cut-off value for the CCPP b-ELISA was set at 50 using 466 CCPP negative and 84 CCPP positive small ruminant sera. Of the negative sera, 307 were obtained from the Botswana National Veterinary Laboratory (BNVL) and 159 from the Friedrich-Loeffler-Institute (FLI) Germany. The 84 positive sera samples came from experimentally vaccinated goats at the AU-PANVAC facility in Debre-Zeit, Ethiopia. The relative diagnostic sensitivity and specificity of the CCPP b-ELISA was 93% and 88%, respectively. This test result indicated good correlation with that of the commercial CCPP cELISA by IDEXX Company (Westbrook, Maine, United States) with a Cohen’s κ agreement of κ agreement of 0.85. The newly developed CCPP b-ELISA will be useful in the detection of antibodies for the diagnosis CCPP and for sero-surveillance during vaccination campaigns.
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