Usutu virus (USUV), one of the most neglected Old World encephalitic flaviviruses, causes epizootics among wild and captive birds and sporadic infection in humans. The dynamics of USUV spread and evolution in its natural hosts are unknown. Here, we present the phylogeny and evolutionary history of all available USUV strains, including 77 newly sequenced complete genomes from a variety of host species at a temporal and spatial scaled resolution. The results showed that USUV can be classified into six distinct lineages and that the most recent common ancestor of the recent European epizootics emerged in Africa at least 500 years ago. We demonstrated that USUV was introduced regularly from Africa into Europe in the last 50 years, and the genetic diversity of European lineages is shaped primarily by in situ evolution, while the African lineages have been driven by extensive gene flow. Most of the amino acid changes are deleterious polymorphisms removed by purifying selection, with adaptive evolution restricted to the NS5 gene and several others evolving under episodic directional selection, indicating that the ecological or immunological factors were mostly the key determinants of USUV dispersal and outbreaks. Host-specific mutations have been detected, while the host transition analysis identified mosquitoes as the most likely origin of the common ancestor and birds as the source of the recent European USUV lineages. Our results suggest that the major migratory bird flyways could predict the continental and intercontinental dispersal patterns of USUV and that migratory birds might act as potential long-distance dispersal vehicles.

Mosquitoes and other arthropods may transmit medically important pathogens, in particular viruses such as West Nile virus. The presence of suitable hosts and competent vectors for those zoonotic viruses is essential for an enzootic transmission, which is a prerequisite for epidemics. To establish reliable risk projections, it is an urgent need for an exact identification of mosquito species, which is especially challenging in the case of sibling species, such as Culex. pipiens pipiens biotypes pipiens and molestus. To facilitate detection of different Culex pipiens forms and their hybrids we established a multiplex real-time PCR. Culex pipiens samples were obtained by egg raft collection and rearing until imago stage or adult sampling using CO2 baited traps and gravid traps. In total, we tested more than 16,500 samples collected all over Germany in the years 2011 and 2012. The predominant species in Germany are Culex pipiens pipiens biotype pipiens and Culex. torrentium, but we also detected Culex pipiens pipiens biotype molestus and hybrids of the two pipiens biotypes at sites where both species occur sympatrically. This report of a potentially important bridge vector for West Nile virus might have major impact in the risk projections for West Nile virus in Germany.

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BackgroundMosquito-borne pathogens are of growing importance in many countries of Europe including Germany. At the same time, the transmission cycles of most mosquito-borne pathogens (e.g. viruses or filarial parasites) are not completely understood. There is especially a lack of knowledge about the vector capacity of the different mosquito species, which is strongly influenced by their host-feeding patterns. While this kind of information is important to identify the relevant vector species, e.g. to direct efficient control measures, studies about the host-feeding patterns of mosquito species in Germany are scarce and outdated.MethodsBetween 2012 and 2015, 775 blood-fed mosquito specimens were collected. Sampling was conducted with Heavy Duty Encephalitis Vector Survey traps, Biogents Sentinel traps, gravid traps, hand-held aspirators, sweep nets, and human-bait collection. The host species for each mosquito specimen was identified with polymerase chain reactions and subsequent Sanger sequencing of the cytochrome b gene.ResultsA total of 32 host species were identified for 23 mosquito species, covering 21 mammalian species (including humans) and eleven bird species. Three mosquito species accounted for nearly three quarters of all collected blood-fed mosquitoes: Aedes vexans (363 specimens, 46.8 % of all mosquito specimens), Culex pipiens pipiens form pipiens (100, 12.9 %) and Ochlerotatus cantans (99, 12.8 %). Non-human mammals dominated the host species (572 specimens, 73.8 % of all mosquito specimens), followed by humans (152, 19.6 %) and birds (51, 6.6 %). The most common host species were roe deer (Capreolus capreolus; 258 mosquito specimens, 33.3 % of all mosquito specimens, 65 % of all mosquito species), humans (Homo sapiens; 152, 19.6 %, 90 %), cattle (Bos taurus; 101, 13.0 %, 60 %), and wild boar (Sus scrofa; 116, 15.0 %, 50 %). There were no statistically significant differences in the spatial-temporal host-feeding patterns of the three most common mosquito species.ConclusionsAlthough the collected blood-fed mosquito species had a strong overlap of host species, two different host-feeding groups were identified with mosquito species feeding on (i) non-human mammals and humans or (ii) birds, non-human mammals, and humans, which make them potential vectors of pathogens only between mammals or between mammals and birds, respectively. Due to the combination of their host-feeding patterns and wide distribution in Germany, Cx. pipiens pipiens form pipiens and Cx. torrentium are potentially most important vectors for pathogens transmitted from birds to humans and the species Ae. vexans for pathogens transmitted from non-human mammals to humans. Finally, the presented study indicated a much broader host range compared to the classifications found in the literature for some of the species, which highlights the need for studies on the host-feeding patterns of mosquitoes to further assess their vector capacity and the disease ecology in Europe.Electronic supplementary materialThe online version of this article (d...

BackgroundTo monitor adult mosquitoes several trapping devices are available. These are differently constructed and use various mechanisms for mosquito attraction, thus resulting in different trapping sensitivities and efficacies for the various species. Mosquito monitoring and surveillance programs in Europe use various types of mosquito traps, but only a few comparisons have been conducted so far. This study compared the performance of four commercial trapping devices, which are commonly used in Europe.MethodsFour different traps, Biogents Sentinel trap (BG trap), Heavy Duty Encephalitis Vector Survey trap (EVS trap), Centres for Disease Control miniature light trap (CDC trap) and Mosquito Magnet Patriot Mosquito trap (MM trap) were compared in a 4 × 4 latin square study. In the years 2012 and 2013, more than seventy 24-hour trap comparisons were conducted at ten different locations in northern and southern Germany, representing urban, forest and floodplain biotopes.ResultsPer 24-hour trapping period, the BG trap caught the widest range of mosquito species, the highest number of individuals of the genus Culex as well as the highest number of individuals of the species Ochlerotatus cantans, Aedes cinereus/geminus, Oc. communis and Culex pipiens/torrentium. The CDC trap revealed best performance for Aedes vexans, whereas the MM trap was most efficient for mosquitoes of the genus Anopheles and the species Oc. geniculatus. The EVS trap did not catch more individuals of any genus or species compared to the other three trapping devices. The BG trap caught the highest number of individuals per trapping period in urban environments as well as in wet forest, while the CDC trap caught the highest number of individuals in the floodplain biotopes. Additionally, the BG trap was most efficient for the number of mosquito species in urban locations.ConclusionThe BG trap showed a significantly better or similar performance compared to the CDC, EVS or MM trap with regard to trapping efficacy for most common mosquito species in Germany, including diversity of mosquito species and number of mosquitoes per trapping period. Thus, the BG trap is probably the best solution for general monitoring or surveillance programs of adult mosquitoes in Central Europe.