BACKGROUND: TAK-003 has been shown to be well tolerated and effective against symptomatic dengue disease and hospitalization, irrespective of baseline serostatus. Most infections are asymptomatic/subclinical. This study assessed whether TAK-003 could protect against asymptomatic/subclinical infections by evaluating increased neutralizing antibody (NAb) titers after natural infection. METHODS: DEN-301 (NCT02747927) is a phase 3 trial among 4- to 16-year-old participants who received 2 doses of TAK-003 or placebo 3 months apart. These exploratory analyses used NAb measured during the trial. As no well-accepted definition for asymptomatic infection exists, 3 algorithms were evaluated: (1) 4-fold increase in NAb, (2) 4-fold increase in NAb and a minimum titer of 40, and (3) 4-fold increase in NAb and a minimum titer of 4-fold lower limit of quantification. Months 4 to 9, months 9 to 15, and months 15 to 27 after first vaccination were analyzed. RESULTS: NAbs from 3765 participants were analyzed. From months 4 to 9, vaccine efficacy (VE) against asymptomatic infection was 51.1% (95% CI, 30.4%-65.6%), 36.1% (95% CI, 6.7%-56.3%), and 27.3% (95% CI, -8.2% to 51.2%) for algorithms 1, 2, and 3, respectively. VE per algorithms 1, 2, and 3 was 54.8% (95% CI, 28.8%-71.3%), 47.9% (95% CI, 16.8%-67.4%), and 44.3% (95% CI, 9.9%-65.6%) in participants with baseline seropositivity and 44.4% (95% CI, 2.1%-68.4%), 4.6% (95% CI, -85.1% to 50.8%), and -29.3% (95% CI, -172.1% to 38.6%) in those with baseline seronegativity. VE against asymptomatic infection gradually decreased from months 4 to 9 to months 9 to 15 and from months 9 to 15 to months 15 to 27. CONCLUSIONS: The variability in VE algorithms indicates challenges in accurately assessing VE against asymptomatic infections. TAK-003 had a modest impact on asymptomatic dengue infections in the first months postvaccination, mainly in participants with baseline seropositivity.