Background

The Asian tiger mosquito, Aedes albopictus, is an aggressive invasive vector responsible for transmitting important arboviruses. Its global spread has been largely facilitated by human-mediated transport, especially through trade and road networks. Since its first detection in the Netherlands in 2005, repeated introductions have occurred via pathways such as lucky bamboo imports, used tires, aircraft, and ground traffic. Despite ongoing surveillance and elimination efforts, uncertainties remain about the origins, recurrence, and establishment potential of these introductions.

Methods

We analyzed 200 Ae. albopictus specimens collected from 21 locations in the Netherlands from 2014 to 2023, including detections in residential areas and points of entry (PoEs). Samples were genotyped at 19 specific microsatellite loci. Genetic diversity and kinship were studied to better understand genetic structure, relatedness between and within locations, and local overwintering. We also determined the presence of Wolbachia endosymbiont strains in the specimens by sequencing Wolbachia markers.

Results

Genetic structure and kinship analyses revealed multiple independent introductions, genetic diversity among sites, and evidence of local overwintering at both residential and PoE locations, including used tire storage sites. Close-kin relationships were identified in 63 specimen dyads. Among these, one dyad confirmed overwintering at a used tire storage facility, four indicated kinship within a residential area, and two between two locations. Genetic assignment results also highlighted successful elimination of the species in one Dutch residential area. A total of 16 Dutch locations (76.19%) tested positive for the presence of Wolbachia. Overall, 48.86% specimens analyzed tested positive for at least one strain, and 35 close-kin dyads showed complete concordance in Wolbachia infection status.

Conclusions

Our findings highlight the complex invasion dynamics of Ae. albopictus in the Netherlands. Our results demonstrate that microsatellite analysis, combined with kinship assessment, is an efficient approach for investigating kinship among individuals within and between urban areas and PoEs, providing evidence of local overwintering, and assessing the genetic structure of Ae. albopictus at introduction sites. The widespread presence of Wolbachia, which is known to reduce mitochondrial diversity, suggests that mitochondrial DNA (mtDNA)-based population analyses may be limited for the species.