(615) Dynamic Fetal T Cell Responses to Viral Exposure Uncovered by Single-cell Systems Immunology

Cytomegalovirus (CMV) is the leading cause of congenital infections, yet the timeline and specificity of fetal adaptive immune responses remain undefined. Prior studies have detected fetal T cell activation in CMV as early as 22 weeks, but lacked patient-level evidence of antigen-specific memory at early gestation. We hypothesized that clonal, CMV-specific fetal T cell memory responses can emerge before 22 weeks gestation and sought to test this by leveraging high-resolution single-cell analyses.

Study Design:

We collected fetal specimens from intrauterine transfusions (IUT; n = 3) and neural tube defect repair (NTD; n = 12) surgeries and used a single-cell multi-omics approach to compare 2 transplacental CMV infections with 12 cases lacking exposure histories. Downstream analyses were conducted with Seurat, clusterProfiler, and scRepertoire for T cell receptor (TCR) profiling.

Results:

Corroborating serological and histopathological clinical diagnoses, CMV gene expression was detected in 26 cells from the two CMV-infected patients, with 52% from monocyte lineages (Fig. 1a). Comparing CMV⁺ and CMV⁻ cells from 166,107 single-cell transcriptomes to a reference of 1,075,352 cells (CZI CellxGene) identified 2,535 differentially expressed genes (DEGs; log2FC >1.4, FDR < 0.05). Pathway analysis revealed 720 significant GO enrichment pathways, highlighted by hundreds of genes in innate, myeloid, and leukocyte pathways (Fig. 1b-c; FDR < 0.05). A focused set of 16,515 cells with paired VDJ sequences revealed 12 unique clusters, including distinct CD8⁺ T cell populations (Fig. 2a). TCR mapping detected clonotypes matching known CMV-specific epitopes (UL29, pp65, IE1; Fig. 2b) and CMV-specific clonal expansion. Gene expression analysis of CMV-specific clonotypes showed limited Th1 responses.
 

Conclusion:

These findings set a new benchmark for fetal immune memory, revealing antigen-specific T cell competence at least three weeks earlier than previously recognized and informing earlier intervention strategies for congenital infections.