Background: The maternally inherited bacteriumWolbachiaoften acts as a subtle parasite that manipulates insect reproduction, resulting potentially in reproductive isolation between host populations. Whilst distinctWolbachiastrains are documented in a group of evolutionarily closely related mosquitoes known as theCulex pipienscomplex, their impact on mosquito population genetics remains unclear. To this aim, we developed a PCR-RFLP test that discriminates the five knownWolbachiagroups found in this host complex. We further examined theWolbachiagenetic diversity, the variability in the coinherited host mitochondria and their partitioning among members of theCx. pipienscomplex, in order to assess the impact ofWolbachiaon host population structure.Results: There was a strong association betweenWolbachiaand mitochondrial haplotypes indicating a stable co-transmission in mosquito populations. Despite evidence that members of theCx. pipienscomplex are genetically distinct on the basis of nuclear DNA, the association ofWolbachiaand mtDNA with members of theCx. pipienscomplex were limited. TheWolbachia wPip-I group, by far the most common, was associated with divergentCx. pipiensmembers, includingCx. quinquefasciatus,Cx. pipiens pipiensformpipiensandCx. pipiens pipiensformmolestus. Four otherwPip groups were also found in mosquito populations and all were shared between diverseCx. pipiensmembers.Conclusion: This data overall supports the hypothesis thatwPip infections, and their allied mitochondria, are associated with regular transfers betweenCx. pipiensmembers rather than specific host associations. Overall, this is suggestive of a recent and likely ongoing cytoplasmic introgression through hybridization events across theCx. pipienscomplex.

Mosquitoes of the Culex pipiens complex are cosmopolitan, and important vectors of neglected tropical diseases, such as arbovirosis and lymphatic filariasis. Among the complex taxa, Cx. pipiens (with two forms pipiens and molestus) and Cx. quinquefasciatus are the most ubiquitous mosquitoes in temperate and tropical regions respectively. Mosquitoes of this taxa lack of morphological differences between females, but have frank behavioral and physiological differences and have different trophic preferences that influence their vectorial status. Hybridization may change the vectorial capacity of these mosquitoes, increasing vector efficiency and medical importance of resulting hybrids. Culex pipiens s.l. from 35 distinct populations were investigated by the study of mtDNA, symbiotic bacterium Wolbachia pipientis, nuclear DNA and flanking region of microsatellite CQ11 polymorphism using PCR with diagnostic primers, RFLP analysis and sequencing. Six different mitochondrial haplotypes were revealed by sequencing of the cytochrome oxidase subunit I (COI) gene and three different Wolbachia (wPip) groups were identified. A strong association was observed between COI haplotypes/groups, wPip groups and taxa; haplogroup A and infection with wPipII appear to be typical for Cx. pipiens form pipiens, haplotype D and infection with wPipIV for form molestus, while haplogroup E, characteristic of Cx. quinquefasciatus, were correlated with wPipI and found in Cx. pipiens sl. from coastal regions of Southern Europe and Mediterranean region. Analysis of microsatellite locus and nuclear DNA revealed hybrids between Cx. pipiens form pipiens and form molestus, as well as between Cx. pipiens and Cx. quinquefasciatus, in Mediterranean populations, as opposed to Northern Europe. Phylogenetic analysis of COI sequences yielded a tree topology that supported the RFLP analysis with significant bootstrap values for haplotype D and haplogroup E. Molecular identification provides the first evidence of the presence of hybrids between Cx. quinquefasciatus and Cx. pipiens as well as cytoplasmic introgression of Cx. quinquefasciatus into Cx. pipiens as a result of hybridization events in coastal regions of Southern Europe and Mediterranean region. Together with observed hybrids between pipiens and molestus forms, these findings point to the presence of hybrids in these areas, with consequent higher potential for disease transmission.

Dirofilariasis is endemic in Russia, as well as in many other European countries. The aim of this study was to assess the ability of mosquitoes to transfer Dirofilaria immitis and Dirofilaria repens in regions with temperate and subtropical climates. The possible impact of the symbiotic bacterium Wolbachia on Dirofilaria transmission was also investigated. 5333 female mosquitoes were collected at 11 points in central European Russia and on the Black Sea coast during the period 2013–2017. Out of 20 mosquito species examined, 14 were infected with D. repens and 13 with D. immitis. Both species of Dirofilaria were found in different climatic regions. The total Dirofilaria spp. estimated infection rate (EIR) in the central part of Russia varied from 3.1% to 3.7% and, in the southern region, from 1.1% to 3.0%. The highest estimated infection rate was found in Anopheles messeae, the lowest in Culex pipiens. The greatest epidemiological danger was represented by Aedes aegypti, Ae. geniculatus, An. messeae and Ae. communis. Six out of 20 mosquito species were infected with Wolbachia. Pools of Aedes albopictus, Cx. pipiens and Coquillettidia richiardii were simultaneously infected with Dirofilaria and Wolbachia. After checking mosquitoes individually, it was found that there was no development of Dirofilaria to the infective larval stage in specimens infected with Wolbachia. Twenty-two Dirofilaria-infective pools were Wolbachia-free and only two mosquito pools were Wolbachia-infected. The potential for transmission of Dirofilaria in mosquito species naturally uninfected with the symbiotic bacterium Wolbachia is higher than in species infected with the bacterium.

To understand specific symbiotic relationships ensuring stable existing of the bacterium Wolbachia in laboratory strains of Drosophila melanogaster, the imago lifespan and senescence rate, as well as

Polymorphism of the gene COI encoding cytochrome C oxidase subunit I and of the rRNA second internal transcribed spacer (ITS2) was measured in Aedes albopictus collected in three locations in Piedmont region and one location in Liguria region in Italy. According to our data, besides the middle part of the COI gene usually used for analysis, there are also SNPs in the 5′ half of the gene, which can provide additional information about the polymorphism of A. albopictus populations. Based on COI gene, three various mitochondrial haplotypes were identified in A. albopictus from four north Italian sampling sites. Different sources of introductions can be considered. The variability between COI sequences of the north Italian A. albopictus is 0.001 while the variability between them based on the pairwise analysis of ITS2 sequences is 10 time more—0.011. For the first time, a 4% divergence between the populations of A. albopictus of tropical and subtropical origins was found on the basis of analyzing the ITS2. A relatively high level of variability of ITS2 between tropical and subtropical populations can be used to develop new markers for phylogenetic studies. No individuals with mitochondrial and nuclear haplotypes specific to tropical populations were found in the four north Italian sampling sites.