Single cell transcriptomes from fetuses exposed to HCMV during gestation reveals a distinct subset of memory-like NKG2C⁺ fetal NK cells

Congenital cytomegalovirus (cCMV) infection is the most common cause of intrauterine infection in the USA, impacting 1 in 100 live births. Although most infected newborns are asymptomatic, 10% display severe congenital anomalies, including microcephaly, sensorineural hearing loss, cerebral palsy, growth restriction, and perinatal mortality. Natural killer (NK) cells are the first lymphocytes to develop during gestation and are required to manage CMV infections. The role of fetal NK cells during cCMV is limited and their development and functions within the umbilical cord has not yet been explored. To investigate this, four sets of umbilical cord blood (UCB) and matching umbilical cord tissues were collected from 2 CMV seropositive (CMV+) and 2 CMV seronegative (CMV-) fetuses that did not experience any complications during gestation. We were able to successfully sort and capture over 5,000 fetal NK cells and perform single-cell RNA-sequencing (scRNA-seq). Using scRNA-seq, we observed and characterized 5 unique fetal NK cell subsets in the UCB and 4 subsets in the corresponding tissue. Interestingly, we identified a distinct subset of NKG2C⁺ memory fetal NK cells displaying elevated expression levels of NKG2C, CD52, CD2, CD16, and CD3E. Here, we demonstrate that cCMV can induce the formation of memory-like NKG2C⁺ NK cells that display a unique transcriptional profile. These findings have the potential to influence the future application of fetal memory NK cells.