Novel Mitochondrial DNA Lineage Found among Ochlerotatus communis (De Geer, 1776) of the Nordic-Baltic Region

Abstract

The Ochlerotatus (Oc.) communis complex consist of three Northern American species as well as a common Holarctic mosquito (Diptera: Culicidae) Oc. communis (De Geer, 1776). These sister species exhibit important ecological differences and are capable of transmitting various pathogens, but cannot always be differentiated by morphological traits. To investigate the Oc. communis complex in Europe, we compared three molecular markers (COI, ND5 and ITS2) from 54 Estonian mosquitoes as well as two COI marker sequences from Sweden. These sequences were subjected to phylogenetic analysis and screened for Wolbachia Hertig and Wolbach symbionts. Within and between groups, distances were calculated for each marker to better understand the relationships among individuals. Results demonstrate that a group of samples, extracted from adult female mosquitoes matching the morphology of Oc. communis, show a marked difference from the main species when comparing the mitochondrial markers COI and ND5. However, there is no variance between the same specimens when considering the nuclear ITS2. We conclude that Oc. communis encompasses two distinct mitochondrial DNA lineages in the Nordic-Baltic region. Further research is needed to investigate the origin and extent of these genetic differences.

Keywords: Ochlerotatus churchillensis; Ochlerotatus nevadensis; Ochlerotatus tahoensis; barcoding; phylogenetics; speciation; vectors.

Figure 1

Map showing the six Estonian collection sites, indicated with green circles. Pie charts demonstrate the amount of Oc. sp. (yellow background) and Oc. communis (orange background) specimen caught from each site. In total this paper includes 20 mosquitoes from site 1 “Undi” (58°29′ N, 26°54′ E), eight from site 2 “Muraste” (59°28′ N, 24°27′ E), four from site 3 “Mändjala” (58°13′ N, 22°20′ E), five from site 4 “Vanajõe” (58°53′ N, 22°26′ E), 16 specimens from site 5 “Metsaküla” and one mosquito from site 6 “Tartu” (58°04′ N, 25°31′ E). 31 of these mosquitoes were caught in 2015 and 23 during 2016. Base map curtesy of ©OpenStreetMap contributors (https://www.openstreetmap.org/copyright) and ©MapTiler (https://www.maptiler.com/copyright/).

Figure 2

Phylogenetic tree based on 72 partial cytochrome c oxidase (COI) sequences (441). Calculated with the Maximum Likelihood method, using Tamura 3-parameter [42] model, with discrete gamma distribution (6 categories (+G, parameter = 0.1790)) Branch lengths are shown to scale and measured as the number of substitutions per site. An. messeae was used as an outgroup for rooting the tree. Oc. sp. sequences are genetically closer to North American species Oc. tahoensis and Oc. churchillensis than the widespread Oc. communis.

Figure 3

Phylogenetic tree representing the information of 48 dehydrogenase subunit 5 (ND5) marker region sequences (321 bp). The tree is calculated with the maximum likelihood method, using the Tamura 3-parameter [42] model and a discrete gamma distribution (6 categories (+G, parameter = 0.1516)). Branch lengths are shown to scale and measured based on the average number of substitutions per site between sequence pairs. An. messeae was used the outgroup in order to root the tree. Oc. sp. sequences cluster together, away from the Oc. communis group.

Figure 4

Phylogenetic tree based on the 53 internal transcribed spacer 2 (ITS2) marker sequences (251 bp). Calculated with the maximum likelihood method, using the Kimura 2-parameter model [43] and discrete Gamma distribution (6 categories (+G, parameter = 0.8737)). Branch lengths are shown to scale and measured based on the average number of substitutions per site between sequence pairs. An. messeae was used as the outgroup for rooting the tree. There appears to be no variation amid the ITS2 sequences Oc. sp. sequences and Oc. communis. However, Oc. churchillensis sequences obtained from GenBank remain as a separate group.