Listeriosis is caused by the bacterium, Listeria monocytogenes, and is a significant health concern because of high hospitalization and mortality rates. This study reports seven cases of pregnancy-associated listeriosis diagnosed with matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry performed on infection-positive blood culture.Blood culture-positive samples of seven patients whose pregnancy was complicated by Listeria infection and treated at Xuanwu Hospital of Capital Medical University between January 2016 and December 2021 were analyzed retrospectively. Strains identified by MALDI-TOF mass spectrometry were compared with colony identification results. Identification accuracy and consistency were assessed.A total of seven strains of Listeria were collected from seven pregnant women presented with fever (37.6-39.9°C). Clinical abnormalities included abnormal liver function, emaciation, hypoalbuminemia, hypocalcemia, hypokalemia, hyponatremia, ketosis, mild to moderate anemia, leukopenia, and thrombocytopenia. Compared with the traditional culture method, MALDI-TOF mass spectrometry led to much earlier identification (4-6 h vs. 3-4 days) with 100% identification accuracy. Of the seven pregnancies complicated by Listeria, only two led to live births. Of the five fetal deaths, three occurred in the second trimester.In this series of pregnancy-associated listeriosis cases, the fetal mortality rate was 71%. MALDI-TOF mass spectrometry is a valuable method that can identify Listeria from blood culture rapidly and accurately.© 2023 Japan Society of Obstetrics and Gynecology.