Key virulence factors responsible for differences in pathogenicity between clinically proven live-attenuated Japanese encephalitis vaccine SA14-14-2 and its pre-attenuated highly virulent parent SA14

• Published in: PLoS pathogens Vol. 21; no. 1; p. e1012844
• Main Authors: Song, Byung-Hak, Yun, Sang-Im, Goldhardt, Joseph L., Kim, Jiyoun, Lee, Young-Min
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science (PLoS) 01.01.2025
• Subjects: Animals; Encephalitis Virus, Japanese - genetics; Encephalitis Virus, Japanese - immunology; Encephalitis Virus, Japanese - pathogenicity; Encephalitis, Japanese - immunology; Encephalitis, Japanese - prevention & control; Encephalitis, Japanese - virology; Female; Humans; Japanese Encephalitis Vaccines - genetics; Japanese Encephalitis Vaccines - immunology; Mice; Vaccines, Attenuated - genetics; Vaccines, Attenuated - immunology; Viral Envelope Proteins - genetics; Viral Envelope Proteins - immunology; Virulence; Virulence Factors - genetics; Virulence Factors - immunology; Virulence Factors - metabolism; Virus Replication
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1012844

Japanese encephalitis virus (JEV), a neuroinvasive and neurovirulent orthoflavivirus, can be prevented in humans with the SA 14 -14-2 vaccine, a live-attenuated version derived from the wild-type SA 14 strain. To determine the viral factors responsible for the differences in pathogenicity between SA 14 and SA 14 -14-2, we initially established a reverse genetics system that includes a pair of full-length infectious cDNAs for both strains. Using this cDNA pair, we then systematically exchanged genomic regions between SA 14 and SA 14 -14-2 to generate 20 chimeric viruses and evaluated their replication capability in cell culture and their pathogenic potential in mice. Our findings revealed the following: ( i ) The single envelope (E) protein of SA 14 -14-2, which contains nine mutations (eight in the ectodomain and one in the stem region), is both necessary and sufficient to render SA 14 non-neuroinvasive and non-neurovirulent. ( ii ) Conversely, the E protein of SA 14 alone is necessary for SA 14 -14-2 to become highly neurovirulent, but it is not sufficient to make it highly neuroinvasive. ( iii ) The limited neuroinvasiveness of an SA 14 -14-2 derivative that contains the E gene of SA 14 significantly increases (approaching that of the wild-type strain) when two viral nonstructural proteins are replaced by their counterparts from SA 14 : ( a ) NS1/1’, which has four mutations on the external surface of the core β-ladder domain; and ( b ) NS2A, which has two mutations in the N-terminal region, including two non-transmembrane α-helices. In line with their roles in viral pathogenicity, the E, NS1/1’, and NS2A genes all contribute to the enhanced spread of the virus in cell culture. Collectively, our data reveal for the first time that the E protein of JEV has a dual function: It is the master regulator of viral neurovirulence and also the primary initiator of viral neuroinvasion. After the initial E-mediated neuroinvasion, the NS1/1’ and NS2A proteins act as secondary promoters, further amplifying viral neuroinvasiveness.