The gobies (Gobiidae) are the most diverse fish family in the Mediterranean Sea. Nevertheless, knowledge on their diversity, taxonomy, and phylogenetic relationships is still inadequate. The phylogenetic analyses reveal two genetically highly distinct clades among specimens identified as Zebrus zebrus. A new species, Zebrus pallaoroi sp. nov., is described based on an integrative approach. The neotype of Zebrus zebrus is designated. Genetic data confirm a pronounced level of divergence between Z. pallaoroi and Z. zebrus, with the mean genetic distance on cytochrome b being 18.1% and 1.07% on rhodopsin. Phylogenetic relationships within the Gobius-lineage were estimated on both markers. Morphologically, Z. pallaoroi is distinguished from the only congener Z. zebrus by having a snout longer than its eye, posterior nostril about 4/5–9/10 of the anterior nostril, eye diameter 4.3−4.7 in head length, ventrolateral head ridges transversally connected on the anterior side by a short transversal ridge, anterior membrane midline depth about 2/3 of the spinous ray, head canal pore α diameter about half of the distance between pore ρ and ρ1, suborbital sensory papillae row 5i going downwards to or near the level of row d, the distance between row 5i and row d absent or much smaller than the length of row 5i, and the body with ten to eleven vertical dark brown bands. Zebrus pallaoroi was recorded from the southern Adriatic, northern Ionian, and northern and western Aegean Seas, and is a cryptobenthic fish from very shallow waters.