Background Several studies have suggested that human cytomegalovirus (CMV) infection is closely related to the pathogenesis of atherosclerosis. The present study aimed to investigate the association between human CMV infection and carotid atherosclerotic plaque vulnerability in a Chinese population. Methods In total, 42 patients with carotid atherosclerosis (observation group) and 30 healthy volunteers (control group) were recruited in our study from October 2016 to January 2018. Statistical analysis was carried out to calculate the infection rate of CMV in subjects. Spearman's rank analysis was performed to evaluate the correlation between CMV infection and atherosclerotic plaque vulnerability. Results The positive rate of CMV was significantly higher in the observation group compared to the control group, and matrix metalloproteinase 9 (MMP-9), tumor necrosis factor-alpha (TNF-alpha) and lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) expression levels were also elevated in the observation group compared to those in the control group. In carotid atherosclerotic patients, the detection rate of unstable plaques and the Crouse scores in vulnerable plaque were significantly higher in the CMV-positive group compared to those in the CMV-negative group. As revealed by correlation analysis, CMV infection was significantly positively correlated with plaque vulnerability and expression levels of MMP-9, TNF-alpha and LOX-1 in carotid atherosclerotic patients. Conclusions Human CMV infection might be a potential risk factor for increased plaque vulnerability in patients with carotid atherosclerosis.