Immune profiling of SARS-CoV-2 infection during pregnancy reveals NK cell and γδ T cell perturbations

**Background:** Pregnant women are considered a vulnerable group for SARS-CoV-2 infection due to physiological and immunological changes during gestation. Studies have associated COVID-19 during pregnancy with increased risk of ICU admission, invasive ventilation, ECMO, death, sepsis, preterm birth, and hypertensive complications. However, comprehensive immunological data comparing pregnant and nonpregnant women with COVID-19 have been lacking. This study aimed to fill this knowledge gap by providing an in-depth analysis of innate, adaptive, and humoral immune responses to SARS-CoV-2 in pregnant women.

**Methods:** The study analyzed 119 women (23 pregnant and 33 nonpregnant with COVID-19; 21 healthy pregnant and 42 healthy nonpregnant controls). Blood samples were collected between March 2020 and March 2021. Acute COVID-19 samples were collected 1-17 days post symptom onset (DPSO), and convalescent samples 21-258 DPSO. The study quantified 217 immunological parameters including: flow cytometry for immune cell activation (NK cells, γδ T cells, MAIT cells, CD4+ and CD8+ T cells, B cells, Tfh cells, monocytes); ELISA for RBD-specific IgG/IgM/IgA and N-specific IgG; surrogate virus neutralization test (sVNT) and microneutralization assays for neutralizing antibodies; a 102-parameter multiplex bead array for systems serology; and a 13-plex cytokine/chemokine panel. Placental tissue from 9 COVID-19 convalescent and 6 unexposed pregnancies was also analyzed.

**Key Results:** - **Antibody responses:** Comparable RBD-specific IgG, IgM, IgA, N-specific IgG titers, and neutralizing activity were observed between pregnant and nonpregnant women with acute or convalescent COVID-19. RBD- and N-specific IgG antibodies crossed the placenta into cord blood, with higher titers in cord blood than matched maternal plasma. - **NK cells:** Healthy pregnant women had significantly higher NK cell activation (HLA-DR+) than nonpregnant women (mean 4.7% vs 0.7%, P<0.001). During acute COVID-19, NK activation in nonpregnant women increased ~11-fold, while in pregnant women it remained at the healthy pregnancy level (fold difference = 1.2). - **γδ T cells:** Healthy pregnant women had higher HLA-DR+CD38+ γδ T cells (mean 4.4% vs 0.8%). Nonpregnant women showed a ~7.5-fold increase during acute COVID-19, while pregnant women showed no further increase above baseline. - **Conventional T cells:** CD4+ and CD8+ T cell activation followed similar trajectories in both groups, with significant increases in HLA-DR+CD38+ CD8+ T cells during acute COVID-19 in both pregnant and nonpregnant women. - **Cytokines:** IL-8, IL-10, and IL-18 were elevated in healthy pregnant women and remained elevated during COVID-19. The IL-18/IL-12p70 ratio was higher in healthy pregnant women, indicating Th2 bias. - **Placental analysis:** Immune cell activation in placentae from COVID-19 convalescent pregnancies was similar to unexposed pregnancies. - **Systems serology:** Principal component analysis revealed distinct antibody and FcγR-binding profiles between pregnant and nonpregnant women, with 27.88% variance explained on PC1 during acute COVID-19.

**Clinical Implications:** This study demonstrates that while adaptive immune responses (antibodies, CD4+ and CD8+ T cells) to SARS-CoV-2 are preserved during pregnancy, the innate immune compartment—particularly NK cells and γδ T cells—shows a preactivated state during healthy pregnancy that prevents further activation during acute infection. This may contribute to the increased vulnerability of pregnant women to severe COVID-19. The preserved antibody responses and placental transfer of IgG provide evidence for vaccine-induced protection in both mothers and neonates. These findings can inform patient management, vaccination strategies, and future research on immune responses during pregnancy.