Detecting bacteria in samples and differentiating between Gram-negative and Gram-positive species is an important challenge, and the most common method, Gram staining, is very time consuming. The aim of this study was to evaluate the electrical bioimpedance spectroscopy (EBIS) technique as an inexpensive and practical tool for real-time detection of bacteria and differentiation between Gram-positive and Gram-negative species. The relevant sensitivity for differentiating between species was found in the magnitude and phase at frequencies of 158,489 and 5248 Hz, respectively, at a bacterial concentration of 1 μg/μl. Subsequently, the sensitivity was estimated as a function of bacterial concentration. Our results demonstrated that EBIS can potentially distinguish between presence and absence of bacteria as well as between different types of bacteria.