Diagnosing infection with small ruminant lentiviruses of genotypes A and B by combining synthetic peptides in ELISA

Abstract

The major challenges in diagnosing small ruminant lentivirus (SRLV) infection include early detection and genotyping of strains of epidemiological interest. A longitudinal study was carried out in Rasa Aragonesa sheep experimentally infected with viral strains of genotypes A or B from Spanish neurological and arthritic SRLV outbreaks, respectively. Sera were tested with two commercial ELISAs, three based on specific peptides and a novel combined peptide ELISA. Three different PCR assays were used to further assess infection status. The kinetics of anti-viral antibody responses were variable, with early diagnosis dependent on the type of ELISA used. Peptide epitopes of SRLV genotypes A and B combined in the same ELISA well enhanced the overall detection rate, whereas single peptides were useful for genotyping the infecting strain (A vs. B). The results of the study suggest that a combined peptide ELISA can be used for serological diagnosis of SRLV infection, with single peptide ELISAs useful for subsequent serotyping.

Phylogenetic Analysis of Belgian Small Ruminant Lentiviruses Supports Cross Species Virus Transmission and Identifies New Subtype B5 Strains

“…Furthermore, sequence information in the gag-pol region, covering epitopes of the capsid protein that are sometimes used as antigens in SRLV ELISAs [24], can also help to identify why some SRLV infected animals are not recognized by currently used ELISA tests and provide the necessary information to construct strain-specific ELISAs using strain-specific epitopes. Proof of concept studies on the use of peptides from SRLV SU5 and transmembrane proteins was already described for the detection of specific Spanish strains that were not detected by other ELISAs [32,33].…”

Post-entry blockade of small ruminant lentiviruses by wild ruminants

“…This was achieved in a first round, by using protein Pierce Purified Recomb ® Protein G Peroxidase conjugated (Recombinant Protein G, Pierce) or secondly, with a secondary antibody able to cross-react with a wide range of ruminant species, including red deer and fallow deer (EG5, Ingenasa). ELISA plates used included commercial Chekit (AG-CHEKIT CAEV/MVV kit, IDEXX Switzerland), ELITEST (Elitest-MVV Hyphen-Biomed, France) and a previously described home-made ELISA that is based on coating with single synthetic peptides alone or in combination [ 39 ]. Procedures were carried out following manufacturer’s instructions with the exception of the conjugate antibody which, as mentioned, was substituted by a secondary antibody able to react with wild species’ IgG.…”

Small Ruminant Lentiviruses in Sheep: Pathology and Tropism of 2 Strains Using the Bone Marrow Route

“…36 In agreement with our ELISA results, previous studies using a peptide-based ELISA on the same animals also showed a higher and more consistent reaction in sheep infected with strain 496 compared with those infected with strain 697. 46 The antigen spectrum covered by a particular test should always be taken into account, and synthetic peptides designed on the basis of circulating strains may improve diagnosis. 12,46 When evaluated clinically, most animals from group B presented carpal enlargement, indicating that the original disease was reproduced experimentally.…”

“…46 The antigen spectrum covered by a particular test should always be taken into account, and synthetic peptides designed on the basis of circulating strains may improve diagnosis. 12,46 When evaluated clinically, most animals from group B presented carpal enlargement, indicating that the original disease was reproduced experimentally. However, this enlargement was not statistically significant, likely because of the difficulty in standardizing the technique employed to measure the carpal joint.…”