Arrival of the Asian tiger mosquito, Aedes albopictus (Skuse, 1895) in Vienna, Austria and initial monitoring activities

Aedes albopictus was recorded in Vienna, Austria, in August 2020 for the first time. The species was found to occur in three sites within the city; morphology-based monitoring was followed by DNA-barcoding. Mitochondrial COI barcode sequences recovered three different haplotypes, however this data does not reveal whether single or multiple introduction events have occurred. The vicinity of Viennese Ae. albopictus sites to major traffic routes highlights the importance of passive transport for range expansion of this species.


INTRODUCTION
In recent decades, several alien mosquito species have been introduced to Europe, primarily through the global transport of goods. In the case of suitable climatic conditions these species were able to form stable populations and further expanded their distribution (Medlock et al., , 2015. Alien mosquito species pose a potential threat, as these newly introduced species may also carry exotic pathogens . Of particular importance is the Asian tiger mosquito (Aedes albopictus), which is not only an annoying day-active biter, but is also able to transmit a broad range of pathogens such as chikungunya, dengue and Zika, which cannot be transmitted by native mosquito species This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2021 The Authors. Transboundary and Emerging Diseases published by Wiley-VCH GmbH in Austria (ECDC, 2021a;Medlock et al., 2012). Aedes albopictus was introduced to Europe mainly with freight transports (especially used tyres and lucky bamboo; Scholte & Schaffner, 2007). From southern Europe, where it has become rapidly established, adults have been passively transported further north in cars and trucks (Eritja et al., 2017;Scholte & Schaffner, 2007). In Germany and Switzerland, this mosquito species has been found in particular along freeway routes from southern Europe (Becker et al., 2013;Flacio et al., 2016;Müller et al., 2020).
Since 2012, single records of Ae. albopictus in southern and western parts of Austria, mostly within a long-term mosquito surveillance program (Fuehrer et al., 2020;Schoener et al., 2019;Seidel et al., 2012), have been limited to small towns and major highway stations, indicative of continuous introduction and local reproduction. At present, there Transbound Emerg Dis. 2021;1-6.
wileyonlinelibrary.com/journal/tbed 1 F I G U R E 1 Location of the first Ae. albopictus findings in Vienna (red boarder), Austria (insert map). Location A -first report by citizen; locations A, B and C lay within the allotment settlement and were investigated during a brief investigation; location V -finding during routine mosquito monitoring program; location MA -reported with Mosquito-Alert app. Grey -major traffic routes is no evidence for overwintering or established populations in Austria.
Here, we report the first finding of this species in Vienna (Austria), one of the largest cities within the European Union.

MATERIAL AND METHODS
On 20 August, 2020, an unusual mosquito specimen was collected by a citizen in an allotment garden in the vicinity of the Danube River in the Leopoldstadt, the second municipal district of Vienna. This citizen had participated in 2018 in a citizen science project whose purpose had been to detect alien mosquito species, using self-made traps in mosquitoalert.com/en), implemented through the AIM-COST Action pan-European surveillance activities (www.aedescost.eu). This specimen was judged by three independent experts to be 'probably Ae.
Following the first report of Ae. albopictus in Leopoldstadt, a brief investigation of the allotment garden was carried out to assess whether this was a stray individual, or if Ae. albopictus was already breeding there. We targeted three locations around the original collection site: location A, the garden of the homeowner who collected the specimen, was monitored with 1 BGS trap baited with carbon dioxide and a specific lure (BG-Sweetscent, Biogents®, Regensburg, Germany) and 2 ovitraps (black 1 L cups filled with ∼750 ml water, with a wooden tongue depressor as oviposition support); location B, ∼600 m from location A and ∼110 m from location C was monitored with an equivalently equipped BGS trap and 2 ovitraps; finally, location C, ∼520 m from location A, was monitored with 2 ovitraps. The traps were run for one week starting on 8 September; the trap nets of the BGS traps were replaced once on 11 September. While the traps were being set up, one Ae. albopictus male was collected with an aspirator.
We performed DNA-barcoding targeting the mitochondrial COI barcode fragment as reported previously (Hebert et al., 2004)  Conventional PCR using primers LepF1 and LepR1 was performed as reported previously (Hebert et al., 2004) and PCR products were sent to LGC Genomics (Berlin, Germany) for bidirectional sequencing.
To investigate the origin of the specimens found DNA haplotype networks were created. NCBI GenBank was mined for COI sequences

RESULTS
In the BGS trap at location A, 111 Ae. albopictus specimens (77 females and 34 males) were collected. In the two ovitraps at this site, 36 and 13 Ae. albopictus eggs were found, respectively. In the BGS trap at location B, no Ae. albopictus were captured. However, in one of the ovitraps at this site, 11 Aedes japonicus (Theobald, 1901) eggs were found. No eggs were found in the ovitraps at location C. In one of the ovitraps Ae. albopictus egg batches belonged to HPT1.

Our genetic analysis on
In the second ovitrap haplotypes HPT1 and HPT2 were found. The adult captured during the mosquito surveillance program in Donaustadt was identified as HPT3.

DISCUSSION
With 1.9 million inhabitants, Vienna is the fifth largest city within the European Union. The many public parks and gardens, and ample suburban areas offer many potential breeding sites, facilitating the establishment of Ae. albopictus populations. In addition, favourable climatic conditions, caused by urban heat-island effects and artificial watering in large cities like Vienna, could increase the survival, breeding success and activity of arthropod vectors such as Ae. albopictus (Bradley & Altizer, 2007). Climate suitability models suggest that the climate in the area of Vienna would permit stable populations of Ae. albopictus to occur, especially if climate warming were to take place (Caminade et al., 2012;Fischer et al., 2011). An establishment of stable populations F I G U R E 2 Median-Joining DNA haplotype network based on a 624 bp section of the mitochondrial COI. The colour-coding of geographic regions follows the United nation geoscheme (see insert). Each coloured circle represents a unique COI haplotype/lineage. The size of the circles corresponds to the frequency, but the number is also indicated for all haplotypes featuring more than two records. Bars on branches indicate the number of substitutions between two haplotypes. Small white circles represent median vectors, which are hypothetical (often ancestral or unsampled) sequences required to connect existing haplotypes with maximum parsimony.

ACKNOWLEDGEMENTS
Financial support for barcoding was provided by the Austrian Federal Ministry of Education, Science and Research via an ABOL (Austrian barcode of Life; www.abol.ac.at) associated project within the framework of the 'Hochschulraum-Strukturmittel' Funds. The preliminary citizen science project was funded by the FWF -Top Citizen Science (TCS-35). The work was done within the framework of AIM-COST Action CA17108. Furthermore, we wish to thank all citizen scientists.

DATA AVAILABILITY STATEMENT
All sequences generated in this study are openly available at the NCBI database (https://www.ncbi.nlm.nih.gov) with the following accession numbers: MW457632-MW457634.