MI2025 – Ruyue Liu

Title:
Broadly cross-reactive B cell clones are derived de novo following sequential orthoflavivirus vaccination

Abstract:
Dengue is an acute mosquito-borne viral disease that poses major global public health concerns, caused by four related but genetically distinct orthoflaviviruses – dengue virus-1 to 4 (DENV-1 to -4). Epidemiological observations have consistently shown that, unlike the first DENV infection that produces type-specific immunity, the second infection elicits broad protection against subsequent infections. However, the B cell receptor (BCR) development following the second compared to the first infection remains unclear. Herein, we leveraged two related live-attenuated orthoflaviviral vaccines, yellow fever (YF17D) and chimeric Japanese encephalitis-yellow fever (JE/YF17D), to simulate sequential orthoflaviviral infections in healthy human volunteers. Participants were first primed with JE/YF17D and then inoculated with YF17D 28 days later; neutralizing antibody titers to both JEV and YF17D were detected at 28 days after YF17D inoculation. FACS sorting, single-cell 3’ reverse transcription, BCR target enrichment, and full-length deep sequencing were carried out on participants’ PBMCs. Even though JEV and YFV share ~70% of the membrane and envelope protein sequence, we found de novo generation of cross-reactive BCR clones after YF17D inoculation. The CDR3 hypervariable region sequences were shared among dominating clones after each dose, but with distinct V(D)J recombination and/or light-heavy chain pairing events, suggesting the insignificance of affinity maturation in sequential vaccination. Skewed V gene usage, preferential V-J pairing, as well as convergent evolution across subjects were also revealed. Various paired, full-length BCR sequences were reconstructed to compare against published orthoflaviviral monoclonal antibody databases. Previously reported Zika virus (ZIKV)-neutralizing antibodies were identified despite clean serological baselines, suggesting strong interspecies protection potential from sequential vaccination. Together, through controlled human infection with live attenuated vaccines, we demonstrated the detailed composition of BCRs with prospective broad-spectrum activity against orthoflaviviruses, which may also serve as a prototype study for human-pathogen interaction research and novel vaccination regime development.

Biography:
Ruyue is a postdoctoral research fellow in Prof. Ooi Eng Eong’s lab at the Programme in Emerging Infectious Diseases, Duke-NUS Medical School. Her research focuses on characterizing immune repertoires in a sequential human infection model, through custom workflows combining single-cell target enrichment and long-read sequencing. She earned her PhD from the National University of Singapore, where she studied Zika virus within-host mutations during the 2016 outbreak in the Americas under the supervision of Dr. October Sessions.