BackgroundInfection is an important risk factor for gestational diabetes mellitus (GDM), while infection of human cytomegalovirus (HCMV) with GDM remains unclear and rarely reported. This study aimed to investigate the association of HCMV infection and serum inflammatory factor levels in pregnancy with GDM.MethodsThis prospective study included pregnant women who attended at Affiliated Hospital of Qingdao Hospital and Zibo Maternal and Child Health Hospital between December 2018 and August 2020. HCMV specific IgM and serum levels of inflammatory factors, including TNF-α, IL-6, and IL-1β, were analyzed.ResultsA total of 5,316 pregnant women were included (415 with GDM (107 with HCMV+GDM+ and 308 with HCMV-GDM+) and 4901 GDM-free (759 with HCMV+GDM- and 4142 with HCMV-GDM-)). The prevalence of GDM was 7.81%. The rate of activation of HCMV was 16.29%. Specifically, 107 and 759 women in the GDM and control group exhibited HCMV infection, with positive rates of25.78% and 15.48%, respectively (P < 0.01). TNF-α, IL-6, and IL-1β at 24–28 weeks of gestation were significantly higher in women with GDM and HCMV infection than inthe other groups (all P < 0.01). Multivariable analysis showed that HCMV positive (OR = 1.851; 95% CI [1.425–2.403]; P < 0.001), IL-6 (OR = 1.010; 95% CI [1.002–1.018]; P = 0.013), and IL-1β (OR = 1.410; 95% CI [1.348–1.474]; P < 0.001) were all significantly correlated with GDM.ConclusionThis study suggests HCMV infection during pregnancy is an independent risk factor of GDM and could significantly increase its incidence. Further studies are needed to elucidate possible mechanisms underlying associations between HCMV infection and GDM.