Context: Physiological fitness testing, such as the Yo-Yo Intermittent recovery test (YYIR) is a key requirement of the Elite Player Performance Plan, introduced by the English Premier League. Eccentric hamstring strength has been identified as a risk factor for hamstring injuries in soccer players, with fatigue highlighted to further exasperate this issue. Objective: The aim of the current study was to examine the effect of the YYIR level 1 (YYIR1) on eccentric knee-flexor strength assessed using the NordBord in youth soccer players. Design: Experimental design. Setting: Soccer club academy. Participants: 67 male academy soccer players (Age = 16.58 0.57 years; Height = 175.45 5.85 cm; Mass = 66.30 8.21 kg) volunteered to participate in the current study during the English competitive soccer season. Main Outcome Measures: Participants conducted eccentric hamstring strength assessments using the NordBord prior to and immediately post completion of the YYIR1, with outcome measures of peak force and peak force relative to body mass recorded. Results: Paired T tests highlighted increased absolute eccentric knee flexor strength values (P < 0.001) immediately post YYIR1 for both the dominant and non-dominant limbs, with the same trend (P < 0.001) observed for eccentric strength relative to body mass. Conclusions: The results of this study indicate that the YYIR1 does not induce eccentric knee-flexor fatigue and as such is not a valid assessment method to assess the effects of fatigue on hamstring function. However, results do suggest that the NordBord may be considered a viable and more accessible alternative to detect pre-post fitness test/fatigue protocol differences in eccentric knee flexor peak strength whilst working in the field.