Early and accurate diagnosis is fundamental for successful surveillance and control of maedi-visna virus (MVV). MVV was detected in Norway in 2019, almost 14 years after the previous outbreak. Genetic analysis indicates persistence of the virus in the sheep population since 2005. The virus was not detected despite continuous serological surveillance. This emphasises the need for improved surveillance, which relies on an understanding of both diagnostic test performance, sampling strategy and the prevalence of the disease. This study therefore aims to evaluate three commercial ELISA tests for MVV antibodies. We conducted a retrospective study using 615 samples from six flocks diagnosed with MVV in 2019. We ran all samples with the following three tests: ID Screen® MVV/CAEV Indirect (IDvet, Grabels, France), IDEXX MVV/CAEV p28 Ab Verification Test (IDEXX Laboratories, Maine, USA) and Elitest MVV/CAEV (Hyphen Biomed, Neuville-sur-Oise, France), hereinafter referred to as ID Screen, IDEXXp28 and Elitest respectively. Without a perfect reference test, we used Bayesian latent class analysis, including conditional dependence between tests, to estimate diagnostic accuracy and true prevalence in the flocks. Using recommended cut-off values, we found that ID Screen and Elitest had significantly higher sensitivity (Se) estimates (99.3 % [97.4-100.0, 95 % Posterior Credible Interval] and 97.4 % [94.1-99.7 %], respectively) than IDEXXp28 (79.5 % [72.3-86.0 %]), while IDEXXp28 and ID Screen had significantly higher specificity (Sp) estimates than Elitest (99.7 % [99.1-100.0], 99.1 % [98.0-99.8 %] and 93.7 % [91.4-95.7 %], respectively). The estimated true prevalence in the six flocks ranged from a median of 0.8-93.5 %. Combining ID Screen and Elitest in serial interpretation showed the highest median Se and Sp (96.7 % [92.0-99.1] and 100.0 % [99.9-100.0], respectively), as well as the highest median positive predictive value (PPV) for the population with the lowest prevalence. Our study supports the use of ID Screen for screening. Further verification with Elitest in serial interpretation will enhance the PPV.

Recent studies that investigated the origins of SRLV strains offered new insights into their distribution among domestic ruminants. The aim of the study was to investigate SRLV circulation in Morocco. A total of 51 farms were selected in different geographical locations and tested by screening and genotyping ELISA. Whole blood was used for DNA extraction and nested gag PCR. The sample size allowed for an estimation of prevalence lower than 20% (CI 95%). Surprisingly, a large proportion of screening-positive samples were not correctly serotyped. Sanger and NGS amplicon sequencing approaches allowed us to obtain new sequences even from difficult-to-amplify samples. The serological data support the evidence of an intrinsic difficulty of SRLV to spread, likely due to management practices. The low rate of success by genotyping ELISA led us to suppose that divergent strains might have escaped from diagnostic tools, as partially confirmed by the evidence of an A subtype carrying a mismatch in serotyping epitope. The sequence analysis revealed the circulation of novel B and recombinant A/B subtypes. This study highlights the importance of monitoring viral sequences and their evolution to develop specific diagnostic tests, particularly in countries where control measures are in place.

Detection of visna/maedi virus (VMV) infection is challenging, as no perfect reference test exists. Several factors can lead to false positive or negative results, including slow seroconversion , fluctuating antibody levels, virus heterogeneity and cross-reactive substances in serum. Norway aims to eliminate VMV, therefore a VMV flock diagnosis will have severe consequences for the farmer. The last two outbreaks occurred in the same geographical area in 2002–2005 and 2019–2020, including 61 and 9 flocks, respectively. The viral strains detected were genetically similar, indicating continuous, undetected VMV circulation in the sheep population. In this descriptive study , we investigate the challenges of correctly diagnosing VMV infection in sheep when the goal is elimination, and describe the distribution of infection within flocks. The choice of test influenced the number of false reactions, as only 40.6 % of the samples that tested positive by the screening ELISA used in 2019–2020 tested positive by two other commercial ELISA tests. In order to minimize the number of false positive VMV flock diagnoses, we investigated if the interpretation could be improved by resampling animals with ambiguous results. Only 26 % of the animals in negative flocks retrieved agreement between the two ELISAs at resampling. It is therefore debatable whether resampling is recommended. Further, we found a higher seroprevalence in older animals, and 36 of 95 offspring obtained different test results than their mother, supporting previous knowledge on slow seroconversion. Different ELISA tests were used in the two outbreaks. Some of the negative samples from 2002 to 2005 were positive when retested with the screening test used in 2019–2020. Hence, animals with false negative results in the 2002–2005 outbreak might be a reason for the continuous circulation of the virus.

Actinobacillus pleuropneumoniae (APP) remains a major cause of respiratory disease in swine and accurate serological tools are essential for surveillance, particularly in sub-clinical herds and low prevalence settings. We evaluated the diagnostic performance of a microsphere-based multiplex fluorescent immunoassay (MFIA) and two commercial ELISAs (1–12 ELISA and ApxIV ELISA) using a Bayesian latent class model and field samples from Norwegian swine herds. Although both ELISAs and the MFIA showed high specificity, the ApxIV ELISA demonstrated lower sensitivity (89.4%) than the other assays. These findings align with previously reported variability in the sensitivity of the ApxIV-based ELISA. The MFIA showed high sensitivity, demonstrating the potential of multiplex immunoassays for APP serology. We also explored optimization of the ApxIV ELISA cut-off, which indicated that lowering the threshold could improve the sensitivity of the test without compromising the specificity for our samples. Our estimates can be influenced by several factors, including the serotype diversity in the population and assumptions related to latent class modelling. Despite these constraints, our findings support the use of MFIA and the ELISAs as accurate tools for APP surveillance, while highlighting that the ApxIV ELISA may be less suitable as a stand-alone screening test. These results underscore the importance of validating diagnostic assays in populations where they will be applied to ensure that they are fit-for-purpose.

The Maedi-visna virus (MVV) causes a persistent infection in small ruminants, and its high genetic heterogeneity affects the performance of diagnostic tests when used in different populations. Therefore, the aim of this study was to develop a bead-based multiplex immunoassay tailored to detect antibodies against a Norwegian MVV strain. We used tissue samples from 14 PCR-positive sheep from a recent MVV outbreak in Norway to sequence the viral strain and produced recombinant antigens based on sequences from one animal. The assay included commercial TM-A and recombinant Norwegian p25, p16–25 and SU5 antigens. Cut-off values for each antigen were determined using receiver operating characteristic curves on 40 ELISA-negative and 67 ELISA-positive samples from the outbreak. The intraplate and interplate repeatability were investigated by testing a quadruplicate of five samples over three days, while the analytical sensitivity (aSe) and specificity (aSp) were measured in comparison to a commercial ELISA. The repeatability showed a coefficient of variation below 15% for most positive samples. The aSe was equal or higher for the multiplex assay than the ELISA, and the aSp of each antigen was 91.7, 93.3, 95.0 and 93.3% for p25, p16–25, SU5 and TM-A, respectively. The assay shows promising results; however, further evaluations of diagnostic characteristics are necessary before implementation in the Norwegian surveillance programme.