The aim of this study was to assess physical fitness, match performance and development of fatigue during competitive matches at two high standards of professional soccer. Computerized time-motion analyses were performed 2-7 times during the competitive season on 18 top-class and 24 moderate professional soccer players. In addition, the players performed the Yo-Yo intermittent recovery test. The top-class players performed 28 and 58% more (P < 0.05) high-intensity running and sprinting, respectively, than the moderate players (2.43 +/- 0.14 vs 1.90 +/- 0.12 km and 0.65 +/- 0.06 vs 0.41 +/- 0.03 km, respectively). The top-class players were better (11%; P < 0.05) on the Yo-Yo intermittent recovery test than the moderate players (2.26 +/- 0.08 vs 2.04 +/- 0.06 km, respectively). The amount of high-intensity running, independent of competitive standard and playing position, was lower (35-45%; P < 0.05) in the last than in the first 15 min of the game. After the 5-min period during which the amount of high-intensity running peaked, performance was reduced (P < 0.05) by 12% in the following 5 min compared with the game average. Substitute players (n = 13) covered 25% more (P < 0.05) ground during the final 15 min of high-intensity running than the other players. The coefficient of variation in high-intensity running was 9.2% between successive matches, whereas it was 24.8% between different stages of the season. Total distance covered and the distance covered in high-intensity running were higher (P < 0.05) for midfield players, full-backs and attackers than for defenders. Attackers and full-backs covered a greater (P < 0.05) distance in sprinting than midfield players and defenders. The midfield players and full-backs covered a greater (P < 0.05) distance than attackers and defenders in the Yo-Yo intermittent recovery test (2.23 +/- 0.10 and 2.21 +/- 0.04 vs 1.99 +/- 0.11 and 1.91 +/- 0.12 km, respectively). The results show that: (1) top-class soccer players performed more high-intensity running during a game and were better at the Yo-Yo test than moderate professional players; (2) fatigue occurred towards the end of matches as well as temporarily during the game, independently of competitive standard and of team position; (3) defenders covered a shorter distance in high-intensity running than players in other playing positions; (4) defenders and attackers had a poorer Yo-Yo intermittent recovery test performance than midfielders and full-backs; and (5) large seasonal changes were observed in physical performance during matches.