Evaluation of Serological Methods and a New Real-Time Nested PCR for Small Ruminant Lentiviruses

Abstract

Small ruminant lentiviruses (SRLVs), i.e., CAEV and MVV, cause insidious infections with life-long persistence and a slowly progressive disease, impairing both animal welfare and productivity in affected herds. The complex diagnosis of SRLVs currently combines serological methods including whole-virus and peptide-based ELISAs and Immunoblot. To improve the current diagnostic protocol, we analyzed 290 sera of animals originating from different European countries in parallel with three commercial screening ELISAs, Immunoblot as a confirmatory assay and five SU5 peptide ELISAs for genotype differentiation. A newly developed nested real-time PCR was carried out for the detection and genotype differentiation of the virus. Using a heat-map display of the combined results, the drawbacks of the current techniques were graphically visualized and quantified. The immunoblot and the SU5-ELISAs exhibited either unsatisfactory sensitivity or insufficient reliability in the differentiation of the causative viral genotype, respectively. The new truth standard was the concordance of the results of two out of three screening ELISAs and the PCR results for serologically false negative samples along with genotype differentiation. Whole-virus antigen-based ELISA showed the highest sensitivity (92.2%) and specificity (98.9%) among the screening tests, whereas PCR exhibited a sensitivity of 75%.

A Combined Approach for Detection of Ovine Small Ruminant Retrovirus Co-Infections

“…Co-infections with different viruses were of low prevalence but did occur [ 32 ], making it more difficult to diagnose them by only one method. Therefore, the use of a combined approach to diagnose these notifiable diseases is recommended.…”

“…Therefore, the use of a combined approach to diagnose these notifiable diseases is recommended. In a recent paper [ 32 ], the combination of ELISA and PCR in serological samples showed co-infection of sheep and goats with SRLV. A co-infection with SRLVs in goats was also described by Pisoni et al [ 47 ].…”

“…Subsequently, a nested real-time PCR using modified primers PI and PVI described by Palmarini et al [ 33 ] and the probe described by Bahari et al [ 36 ] was performed ( Figure 1 ). The amplification steps for the exJSRV-PCR nested real-time PCR and the nested real-time PCRs for CAEV and MVV were carried out as described by Schaer et al [ 32 ]. The exJSRV real-time PCR for paraffin-embedded tissue samples was validated with tissue samples of 40 histopathological confirmed cases of OPA.…”

“…The ongoing SRLV diagnostic strategy in Switzerland is based on an array of four sequentially used serological tests [ 31 ] on single animals. Recently, a nested real-time PCR protocol with the ability to differentiate between CAEV and MVV was developed [ 32 ]. Confirmation of the disease in basic necropsy protocols in the pathology field requires the detection of the causative viruses, optimally both at molecular and in situ level.…”

Longitudinal study on seroreactivity of goats exposed to colostrum and milk of small ruminant lentivirus–infected dams

“…Genotypes of SRLV have been shown to vary in terms of their potential for lactogenic transmission (51) as well as antigenic properties (11,22,45). As a result, the diagnostic performance of serological assays varies depending on the degree of sameness of the genotype responsible for the infection and the genotype from which the antigens used in the assay were derived (15,33,42,52,58,60). Although very few studies have investigated the differences in antibody response between goats infected with genotype A and those infected with genotype B, the humoral response after infection with the homologous SRLV genotype has been shown to be stronger than that which followed the heterologous genotype infection in both goats and sheep (33,41).…”