The expansion of invasive alien species is considered a major threat to forest ecosystems and biodiversity. Their potential impacts range from local changes in species composition to wider-scale effects on forest habitat and landscape functioning, although the latter has been relatively little explored in the literature. Here, we assessed the impact of an invasive forest pest, the pine wood nematode (PWN), in the Natura 2000 network of protected areas (PAs) in Portugal, the first European country in which PWN was reported. We considered the impacts of the pest’s spread (up to 2016) on individual PAs, in terms of the fraction of their coniferous forest infected, and on the corridors between PAs, which were mapped and prioritized through least-cost path modelling, geographic information system analysis, and the graph-based probability of connectivity metric. We found that PWN by 2016 had spread into 49% of the Portuguese Natura 2000 coniferous forest habitat, while it had invaded 68% of the coniferous forests that form the priority corridors between the PAs. These impacts are likely to be aggravated in the next years, given the pace of PWN expansion and the predicted rates of natural spread to new areas in Portugal and, increasingly likely, in Spain. Our results suggest that the connectivity of PA systems may be significantly disrupted by alien species, and that spatially prioritized control measures can help mitigate the impacts of invasive species on the coherence and functionality of protected area networks such as Natura 2000.