This study aimed to investigate the effects of online physical education classes, using Tabata training, on middle school students’ physical fitness. Fifty-four adolescents were randomly assigned to either the asynchronous online class group (AOCG, n = 24, age: 15.8 ± 0.4 years) or the synchronous online class group (SOCG, n = 24 age: 15.9 ± 0.3 years). The online physical education class lasted two days per week for 10 weeks. Recorded video lectures were conducted for the AOCG, and Tabata training for the SOCG, as real-time lecture methods. Baseline and post-online physical education class measures included muscular strength, muscular endurance, flexibility, balance, and cardiorespiratory fitness tests. The results showed that the synchronous online physical education class had a positive effect on the improvement of muscle mass, ankle strength (dorsiflexion), hip strength (abduction, flexion, extension, and external rotation), knee strength (extension and flexion), and balance (Y-balance test) in adolescents. These findings suggest that the physical fitness of adolescents can be sufficiently improved through appropriate online physical education class methods. Further research should focus on developing and evaluating different types of exercises for synchronous online physical education classes as a precautionary measure for the second wave of COVID-19.