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Background: We aimed to investigate the effects of incremental PEEP titration in patients with ARDS on regional diaphragmatic motion with bedside ultrasound., Methods: Dorsal diaphragmatic excursion (DDE) and ventral diaphragmatic excursion (VDE) were measured using anatomic M-mode ultrasonography of the right hemidiaphragm as PEEP was randomized to -6, -3, +3, and +6 cm H 2 O from baseline to achieve a positive transpulmonary pressure. Inter-operator variability of DDE was assessed in 10 separate subjects., Results: A total of 14 subjects ventilated for ARDS were enrolled. Subjects had a mean age of 54 ± 12 y, mean P aO 2 /F IO 2 = 137 ± 54 mm Hg, and mean sequential organ failure assessment (SOFA) score = 14 ± 1). Transpulmonary pressure, DDE, and DDE/VDE ratio increased with incremental PEEP titration (-1.15 cm H 2 O vs 3.63 cm H 2 O, P < .001; 4.9 mm vs 8.2 mm, P < .001; and 62% vs 93%, P < .001, respectively). When transpulmonary pressure became positive, a visual increase in DDE and DDE/VDE ratio 0.60 to 0.93 was observed (from 0.48 cm to 0.82 cm, R 2 = 0.87, P = .02; and R 2 = 0.93, P = .006, respectively). There was high agreement in DDE measurements between 2 ultrasonographers (intra-class correlation 0.987, P < .001)., Conclusions: DDE was affected by incremental PEEP titration toward a positive transpulmonary pressure. The ultrasound assessment using anatomic M-mode allowed for specific measurement of regional diaphragmatic excursion. This pattern of motion in the dependent regions of the diaphragm during PEEP titration in subjects with ARDS achieving a positive transpulmonary pressure may reflect a potential target for future studies in the bedside assessment for lung recruitment. (Clinical Trials.gov registration NCT02463773.)., Competing Interests: The authors have disclosed no conflicts of interest., (Copyright © 2020 by Daedalus Enterprises.)