Developing a detection method for rabies virus neutralizing antibodies using pseudotype technique

Abstract

Rabies is a neuro-fatal disease, causing by rabies virus (RABV) infection. Two serological tests, recommended by the World Health Organization and the World Organization for Animal Health, namely the rapid fluorescent focus inhibition test (RFFIT) and the fluorescent antibody virus neutralization test (FAVN) are considered gold standard. However, as both the RFFIT and FAVN require the use of live viruses, they raise biosafety concerns. Moreover, the immunostaining step in both methods is costly and time-consuming. In this study, RABV-pseudotype was developed and used in a RVNA detection method. The RABV-pseudotype based on lentivirus gave higher titer than the vesicular stomatitis virus (VSV). Fifty dog serum samples were tested for RVNA titer and compared with FAVN to validate the new pseudotypre-based method. The diagnostic sensitivity and specificity of this method was 92% and 100%, respectively. The analytical specificity of the test was confirmed by lacking of cross-neutralization with an anti-CDV monoclonal antibody. The test repeatability was demonstrated by the coefficient of variation of 1.33 among 4 different timepoints. The RVNA titer measured by both methods was in a strong positive correlation (Pearson r = 0.9491, p < 0.0001). In conclusion, the RABV pseudotype-based assay developed in this study offers a safer and faster means for assessing the immune status of the dog population.