Background Aedes japonicus japonicus has expanded beyond its native range and has established in multiple European countries, including Belgium. In addition to the population located at Natoye, Belgium, locally established since 2002, specimens were recently collected along the Belgian border. The first objective of this study was therefore to investigate the origin of these new introductions, which were assumed to be related to the expansion of the nearby population in western Germany. Also, an intensive elimination campaign was undertaken at Natoye between 2012 and 2015, after which the species was declared to be eradicated. This species was re-detected in 2017, and thus the second objective was to investigate if these specimens resulted from a new introduction event and/or from a few undetected specimens that escaped the elimination campaign. Methods Population genetic variation at nad4 and seven microsatellite loci was surveyed in 224 and 68 specimens collected in Belgium and Germany, respectively. German samples were included as reference to investigate putative introduction source(s). At Natoye, 52 and 135 specimens were collected before and after the elimination campaign, respectively, to investigate temporal changes in the genetic composition and diversity. Results At Natoye, the genotypic microsatellite make-up showed a clear difference before and after the elimination campaign. Also, the population after 2017 displayed an increased allelic richness and number of private alleles, indicative of new introduction(s). However, the Natoye population present before the elimination programme is believed to have survived at low density. At the Belgian border, clustering results suggest a relation with the western German population. Whether the introduction(s) occur via passive human-mediated ground transport or, alternatively, by natural spread cannot be determined yet from the dataset. Conclusion Further introductions within Belgium are expected to occur in the near future, especially along the eastern Belgian border, which is at the front of the invasion of Ae. japonicus towards the west. Our results also point to the complexity of controlling invasive species, since 4 years of intense control measures were found to be not completely successful at eliminating this exotic at Natoye.

The present study aimed at identifying the members of the Anopheles maculipennis complex (Diptera: Culicidae) occurring in Belgium. Therefore, the second internal transcribed spacer of nuclear ribosomal DNA (ITS2) and the mitochondrial cytochrome oxidase subunit I (COI) loci were sequenced in 175 and 111 specimens, respectively, collected between 2007 and 2019. In parallel, the suitability of two species‐diagnostic PCR‐RFLP assays was tested. The identified specimens included: An. maculipennis s.s. (N = 105), An. daciae (N = 62), An. atroparvus (N = 6) and An. messeae (N = 2). Each species was characterized by unique ITS2 haplotypes, whereas COI only supported the monophyly of An. atroparvus, a historical malaria vector in Belgium. Species identification results were further supported by unique PCR‐RFLP banding patterns. We report for the first time An. daciae in Belgium, where it was found to co‐occur with An. maculipennis s.s. The latter was the most prevalent in the collection studied (60%) and appears to have the widest distribution in Belgium. As in other studies, An. daciae and An. messeae appeared the most closely related species, up to the point that their species status remains debatable, while their ecological differences, including vector competences, need further study.

Many symbionts live in association with different species. It can be expected that these distinct hosts might have a different effect on key life-history traits of the associated symbionts. Here, we compared the key trait body size of the obligatorily ant-associated isopod Platyarthrus hoffmannseggii collected in nests of two types of sympatric ant hosts. This isopod species showed surprisingly large differences in body size depending on type of host ant, with the head width of females and males associated with organic mound building red wood ants (RWAs) being, respectively, 1.30 and 1.17 times larger than isopods sympatrically living in earth nests of Lasius flavus. There was also a higher proportion of females in many RWA nests, but this pattern was not consistent across all nests. Genetic analyses and aggression trials did not reveal cryptic groups specialized to different hosts. Therefore, we argue that the isopods exhibit size plasticity because of different host nest conditions. Absence of host aggression and optimal abiotic conditions in RWA nests might promote a larger isopod body size. Overall, this study shows that the association of a symbiont with different hosts might induce phenotypic plasticity in a symbiont key trait.

This survey reports on the DNA identification and occurrence of Culex torrentium and Cx. pipiens s.s. in Belgium. These native disease-vector mosquito species are morphologically difficult to separate, and the biotypes of Cx. pipiens s.s. are morphologically indistinguishable. Culex torrentium and Cx. pipiens s.s. were identified using the COI and ACE2 loci. We recorded 1248 Cx. pipiens s.s. and 401 Cx. torrentium specimens from 24 locations in Belgium (collected between 2017 and 2019). Culex pipiens biotypes pipiens and molestus, and their hybrids, were differentiated using fragment-size analysis of the CQ11 locus (956 pipiens and 227 molestus biotype specimens, 29 hybrids). Hybrids were observed at 13 out of 16 sympatric sites. These results confirm that both species are widespread in Belgium, but while Cx. torrentium revealed many COI haplotypes, Cx. pipiens s.s. showed only one abundant haplotype. This latter observation may either reflect a recent population-wide demographic or range expansion, or a recent bottleneck, possibly linked to a Wolbachia infection. Finally, new evidence is provided for the asymmetric but limited introgression of the molestus biotype into the pipiens biotype.