The coloNISation: spatio-temporal metabarcoding surveys in ports reveal homogenised communities with high genetic diversity and connectivity of non-indigenous species

Large commercial ports facilitate the introduction of non-indigenous species (NIS), while smaller harbours and marinas contribute to their regional spread. Harbour networks are thus important drivers of introductions. Despite extensive research effort on NIS in recent years, no study has yet assessed genetic connectivity among harbours considering whole-community composition. Here, we analysed spatio-temporal patterns of metazoan communities over one year in four medium-size harbours along the NW Mediterranean coast sampled by deploying standardised biological collectors. Using cytochrome c oxidase subunit I (COI) metabarcoding, we identified 1,770 metazoan molecular operational taxonomic units (MOTUs), of which 82 were classified as NIS based on a custom database of Mediterranean NIS. Despite their lower species count compared to natives, NIS accounted for 34–70% of reads in harbours. The southernmost harbour had the highest NIS number of reads, likely due to its proximity to aquaculture facilities. While we observed some variation in the spatial structure of metazoan communities across harbours, NIS showed consistently low differentiation values, sharing significantly more MOTUs among sites. Seasonal patterns influenced both NIS and the rest of the community. Haplotype diversity was significantly higher in NIS, which also exhibited lower genetic differentiation across harbours compared to native species, indicating NIS spread via local boating and likely recurrent introductions. These findings highlight distinct dynamics between NIS and native species in artificial environments, emphasising the importance of continued monitoring in harbour networks to manage coastal NIS proliferation.