Epitope mapping of diagnostic anti-rabies monoclonal antibodies provide structural insight into rabies variant identification in infected wildlife.

Abstract

Rabies virus is a member of the lyssavirus family and causes over 60,000 deaths a year. There are currently five strains of rabies virus circulating throughout the continental United States. In order to better control the spread of rabies virus, the Center of Disease Control monitors the prevalence and host range of different rabies virus variants (RVV) throughout the United States. This surveillance relies on a panel of monoclonal antibodies that are known to differentially bind to the nucleoprotein of RVV in a direct fluorescent antibody test (DFA). While these tests are known to accurately distinguish RVV, it is unknown where these antibodies bind to the rabies nucleoprotein. To better understand the fundamental principles of this assay, the epitopes of three monoclonal antibodies from the current panel, which are suspected to have conformational epitopes, were determined using hydrogen-deuterium mass spectrometry. We determined these antibodies bind to distinct surface-exposed epitopes of the nucleoprotein, one of which makes contacts with multiple subunits of the 11-mer ring that is formed by the nucleoprotein. After determining the epitopes, a multiple sequence alignment was examined to look at the conservation of the amino acids in each epitope to determine how these antibodies distinguish between RVV. By determining epitopes, we can now predict how viral mutations could change the specificity and/or sensitivity of a rabies DFA. The results of this work can be used to help further develop rapid immunoassays for more economical RVV diagnostics in the field, in regions that may lack access to microscopy-based DFA diagnostics.