To assess the relationship between the absence of vaginal lactobacilli and preterm birth at 37 weeks of gestation. Lactobacilli (odds ratio and 95% confidence interval: 0.15 [0.09 to 0.24]), Mycoplasma hominis (2.3 [1.0 to 5.4]), and glucose non-fermentative gram-negative rods (2.1 [1.0 to 4.2]) were identified as independent risk factors for preterm delivery at < 33 weeks of gestation. Absence of lactobacilli (sensitivity and positive predictive value: 28% and 25%) was a better predictor of preterm delivery at < 33 weeks of gestation than the presence of Mycoplasma hominis (7% and 13%, respectively) or glucose non-fermentative rods (9% and 11%).Although this was not a cohort study, results suggest that tests for determining the presence of vaginal lactobacilli may be clinically useful tools for identifying women at an increased risk of preterm delivery at < 33 weeks of gestation.