Development and validation of an indirect ELISA for the serodiagnosis of contagious caprine pleuropneumonia in goats

Abstract

Mycoplasma capricolum subsp. capripneumoniae (Mccp) is a dreadful pathogen of contagious caprine pleuropneumonia (CCPP). Mccp local strains have recently been isolated from naturally infected goats in northern areas of Pakistan. In this study, we established an in-house indirect ELISA (iELISA) based on whole-cell proteins (WCP) of local Mccp strains for the detection of anti-Mccp antibodies. Serum samples from naturally (n=500) and experimentally infected (n=20) and non-infected animals (n=100) were used for the establishment of iELISA. The specificity and sensitivity of the iELISA were determined using a commercially available competitive ELISA (cELISA) kit as a reference standard. The cut-off value (OD₄₅₀) was found 0.53 by ROC analysis. As indicated by a kappa value of 0.88 and 96.66% agreement, both assays showed perfect agreement. An iELISA based on WCP of local Mccp detected seroconversion in experimental goats 7 days post-infection (dpi). The iELISA revealed a sensitivity of 90.83% (95% CI: 83.77%, 95.55%) and a specificity of 92.31% (95% CI: 84.79%, 96.85%) and has no cross-reaction with related mycoplasma epidemic diseases. For both inter-assay and intra-assay precision, the iELISA percentage coefficient of variation (%CV) was 4.8% and 1.74%, respectively. In conclusion, this in-house iELISA based on local Mccp-WCP was determined as a sensitive and specific assay for the diagnosis of Mccp infection.