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Molecular characterization of attenuated Japanese encephalitis live vaccine strain ML-17
Paresh Sumatilal Shah1, Mariko Tanaka, Afjal Hossain Khan
1Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan.
Abstract:
The Japanese encephalitis (JE) zoonotic vaccine strain ML-17 was sequenced and compared to related JE virus strains to identify genomic attenuation markers. Relative to its parental strain, JaOH0566, 25 nucleotide alterations and 10 amino acid changes to, prM/M(2), NS2A(1), NS4B(3) and NS5(4) proteins were recorded. Both structural-gene changes were in the prM/M region (127Met-->Ile and 274Asn-->Thr). To study the effects of these prM/M changes, mutants bearing the changes were prepared using an infectious clone of JaOArS982 previously established at this lab. Compared with JaOArS982, mutant 127(Met-->Ile) showed marked reduction in murine neuroinvasiveness. Mutant 274(Asn-->Thr), showed slight reduction. Neither mutant recorded ML-17-equivalent attenuation, implying that prM/M changes need to combine with other recorded genomic differences to cause attenuation. Importantly, ML-17 with its unchanged E region, presents a possible backbone candidate for preparation of "E-replacement" type live attenuated flavivirus chimeric vaccines.
Insights
Japanese encephalitis (JE) vaccine strain ML-17 has genomic differences from its parent strain. Specific prM/M protein changes reduce neuroinvasiveness, suggesting combined genomic alterations are key for attenuation.
Area of Science:
- Virology
- Genomics
- Vaccine Development
Background:
- Japanese encephalitis (JE) is a significant mosquito-borne viral disease.
- Live attenuated vaccines are crucial for JE control.
- Identifying genomic markers of attenuation is vital for vaccine improvement.
Purpose of the Study:
- To sequence and compare the JE vaccine strain ML-17 with related strains.
- To identify genomic attenuation markers within the ML-17 strain.
- To investigate the role of prM/M protein changes in viral attenuation.
Main Methods:
- Whole-genome sequencing of JE virus strain ML-17.
- Comparative genomic analysis with parental and related JE virus strains.
- Site-directed mutagenesis using infectious clones to create prM/M mutants.
- Assessment of neuroinvasiveness in murine models.
Main Results:
- ML-17 exhibited 25 nucleotide and 10 amino acid alterations compared to its parental strain JaOH0566.
- Two specific amino acid changes in the prM/M protein (Met127Ile and Asn274Thr) were identified.
- Mutant 127(Met-->Ile) showed a marked reduction in murine neuroinvasiveness, while mutant 274(Asn-->Thr) showed a slight reduction.
- Neither single prM/M mutation conferred ML-17-equivalent attenuation, indicating a need for combined genomic changes.
Conclusions:
- prM/M protein changes contribute to JE virus attenuation but require combination with other genomic differences.
- The JE vaccine strain ML-17, with an unchanged E protein region, is a potential candidate for developing "E-replacement" chimeric flavivirus vaccines.
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