The hidden invader: ascidian Didemnum pseudovexillum expands eastward in two Mediterranean non-indigenous species hotspots

Biological invasions are escalating threats to marine ecosystems, and cryptic and/or cryptogenic species are especially problematic because their morphological similarity to native or established taxa may lead to undetected introductions and delayed management responses. Among these taxa, colonial ascidians are widespread biofouling organisms on artificial infrastructures, frequently introduced and challenging to identify. Here we document the establishment of the ascidian Didemnum pseudovexillum Turon and Viard, 2020 in two Eastern Mediterranean hotspots of non-indigenous species (NIS), the Venice Lagoon and Gulf of Taranto. Integrative taxonomy based on morphological and COI-based analyses confirmed species identity and revealed persistent colonization of artificial substrates, with larvae recorded in autumn. We characterized the environmental niche of D. pseudovexillum across local temperature–salinity gradients and its reproductive window in the northern Adriatic and Ionian Seas. To evaluate the wider invasion threat posed by D. vexillum and D. pseudovexillum, we developed a multi-factor risk assessment for 21 Mediterranean ports and aquaculture hubs. Environmental suitability, shipping connectivity, diversity of established NIS ascidians, and aquaculture intensity were combined into a composite risk index, and its sensitivity to alternative weighting schemes was explored. Our results highlight D. pseudovexillum as an overlooked invader in the whole Mediterranean and provide an operational basis for prioritizing molecular surveillance of this species and other cryptic ascidians in NIS hotspots.