Crosstalk is an important source of error in interpreting surface electromyography (EMG) signals. Here, we aimed at characterizing crosstalk for three groups of synergistic muscles by the identification of individual motor unit action potentials. Moreover, we explored whether spatial filtering (single and double differential) of the EMG signals influences the level of crosstalk. Three experiments were conducted. Participants (total twenty-five) performed isometric contractions at 10% of the maximal voluntary contraction (MVC) with digit muscles and knee extensors, and at 30% MVC with plantar flexors. High-density surface EMG signals were recorded and decomposed into motor unit spike trains. For each muscle, we quantified the crosstalk induced to neighboring muscles and the level of contamination by the nearby muscle activity. We also estimated the influence of crosstalk on the EMG power spectrum and intermuscular correlation. Most motor units (80%) generated significant crosstalk signals to neighboring muscle EMG in monopolar recording mode, but this proportion decreased with spatial filtering (50% and 42% for single and double differential, respectively). Crosstalk induced overestimations of intermuscular correlation and has a small effect on the EMG power spectrum, which indicates that crosstalk is not reduced with high-pass temporal filtering. Conversely, spatial filtering diminished the crosstalk magnitude and the overestimations of intermuscular correlation, confirming to be an effective and simple technique to reduce crosstalk. This paper presents a new method for the identification and quantification of crosstalk at the motor unit level and clarifies the influence of crosstalk on EMG interpretation for muscles with different anatomy.