JOURNAL OF ENTOMOLOGY EUROPEAN JOURNAL OF ENTOMOLOGY The northward spread of the European mantis, Mantis religiosa (Mantodea: Mantidae): Data from Lithuania

. Geographic distribution of the European mantis, Mantis religiosa (Linnaeus, 1758) in Europe covers mostly southern and central Europe, but this species has recently shown a signi ﬁ cant northward spread. First reports of M. religiosa in Lithuania were in 2008 and now these insects are distributed throughout this country. Information on the spread of M. religiosa in Lithuania between 2015 and 2020 are analysed in this paper. The spread of this insect in Lithuania and neighbouring countries can be related to changes in climate, in particular, the increase in average annual temperature and milder winters . Possible routes along which M. religiosa spread northwards based on an analysis of mtDNA sequences are presented.

Increase in economic activity and combustion of fossil fuels over the last century greatly affected the environment, which can be seen in the change in climate. The average global land and ocean surface temperatures have increased linearly by 0.85°C over the period 1880 to 2012 ( Pachauri & Meyer, 2014). The year 2008 was the warmest in Lithuania over the period 1778 to 2010, with an average annual temperature of 8.3°C and positive average annual anomaly of 2. 1°C (Galvonaite et al., 2013). These climate changes have already affected many natural systems in terms of the timing of seasonal biological events and geographic shifts in the ranges of species ( Semenza & Menne, 2009).
During the last few years M. religiosa was reported in Lithuania and some even more northerly countries such as Latvia and Estonia for the fi rst time ( Pupiņš et al., 2012;Truuverk, 2019). The presence of M. religiosa in Lithuania was fi rst reported in 2008 and is now distributed almost throughout the country. Here we present data from Lithuania on the dispersal of these insects over the period 2015-2020. P ossible sources of M. religiosa based on an analysis of COI sequences of the specimens collected are also presented. This data may be useful for predicting the spread of other insect species, especially in regard to climate change.

Insect collection and identifi cation
The information on the detection of M. religiosa in Lithuania in 2015-2020 was collected using an informative campaign and

INTRODUCTION
Insects of the order Mantodea are mostly distributed in tropical areas and only a few occur in the warmer parts of the temperate zone ( Kočárek et al., 2005). These insects are predators and are characterized by raptorial fore legs adapted for seizing prey, the mid and hind legs being cursorial.
During the fi ve-year investigation (2015-2020) 433 confi rmed records of the presence of M. religiosa in different parts of Lithuania were reported based on a total of 481 specimens (adults and juvenile), as in some cases several individual insects were recorded for the same locality. Mantis religiosa were recorded in 33 different districts and municipalities.
Numbers of districts in which insects were recorded each year varied between 11 (in 2016) and 22 (in 2020), and the number of M. religiosa recorded per year was highest in 2020 ( Fig. 1). Most were recorded in the east (41.4%) and south (25.8%) of Lithuania, with 12.8% and 11.8% recorded in central and south-eastern parts, and only a few in the northwest (0.2%), west (1.4%), north (1.6%,), southwest (2.3%) and northeast (2.7%). Until the year 2017 M. religiosa were recorded only from eastern, north-eastern, southern and central Lithuania with most reports for the districts Vilnius and Kaunas (Fig. 2). Since 2018, the number of reports from central Lithuania has increased, with records of M. religiosa in Šiauliai district in the north and Jurbarkas district in the west. After 2019 insects were recorded in the west, reaching Pagėgiai district and the Curonian spit on the Baltic Sea in the very west of Lithuania (Fig. 2). Females accounted for 52.5%, males 22.6% and immature individuals 3.8% of all specimens of M. religiosa recorded, 21.1% of which the sex was not determined. Mantis questioning inhabitants (389 records), and collecting or observing insects during fi eld studies by the authors in 2008-2020. Field studies included using light traps, observation of habitats at night-time using a head lamp and scanning vegetation in daytime. Mantis religiosa has a distinctive appearance, are attracted to city lights at night and are familiar and recognizable, even by non-professionals. Thus, it was possible to use a questionnaire to determine the distribution of these insects. The fi rst information on the presence of this insect aroused the interest of the media and citizens generally, which enabled us to use newspapers, the internet, radio and television to obtain information from the inhabitants of Lithuania. The media requested people to report information on this insect, such as the place seen, habitat, date and a photograph. Additional information was obtained by direct communication. The data (44 records) in the most popular online database the social network for naturalists, "iNaturalist" (available from https://www.inaturalist.org, accessed 06 05 2021) were also used. The announcer passed on the captured individuals in some cases. Specimens were identifi ed using the keys of Bey-Bienko (1964), Kočárek et al. (2005) and Shc herbakov et al. (2015). The collected material is deposited in Tadas Ivanauskas Zoological Museum (Kaunas, Lithuania) and in the entomological collections of the Nature Research Centre (Vilnius, Lithuania).
Sixteen insects collected in Lithuania in 2015-2019 and parts of legs of specimens collected in Czech Republic (1 spec.), Belarus (1 spec.) and ootheca (1) from France were preserved in 96% ethanol for the PCR-based investigation.
All reactions were done in a 25-μL volume for each sample. It consisted of 12.5 μL DreamTag Master Mix (Thermofi sher, Lithuania), 8.5 μL nuclease-free water, 1 μL of each primer and 2 μL of template DNA. The amplifi cation was evaluated by running 2 μL of each PCR product on 2% agarose gel. Fragments were sequenced of both strains with corresponding primers and with the BigDye Terminator V3.1 Cycle Sequencing kit according to the manufacturer's instructions in the Nature Research Centre. Sequences were edited and aligned using the program BioEdit (Hall, 1999). Phy logenetic tre es were produced, and intraspecifi c genetic distances were based on the Kimura 2-Parameter model and calculated using MEGA software.

Meteorological variables
Data on the mean annual air temperature (°C), mean January air temperature (°C) and air temperature anomalies compared to climatic norm in Lithuania were obtained from the archive of t he Lithuanian H ydrometeorological Service. January air temperature was used because the coldest month in Lithuania is usually January (Galvonaite et al ., 2013). The average air temperature religiosa of different colours were recorded in Lithuania during the period of this study: green (90.8%) and light brown (9.2%). Of the brown individuals most were female (79.5%), with 15.9% male and 4.5% immature. Of the green individuals, most were female (66.2%), with 28.9% male and 4.9% immature. There was no signifi cant difference in the sex ratio of mantids of the different colours (p = 0.06, χ 2 = 3.47).
The earliest date on which adults of M. religiosa were observed was the 29 th of July (2020) and the latest the 25 th of October (2020), they were most abundant (peaks) in August in all years of this study, but 2016 (the fi rst decade of October) and 2017 (the third decade of September) (Fig.  3). Periods when the peaks of M. religiosa were recorded in August differed slightly each year and were recorded during the fi rst (2015), the second (2018, 2019) and the third decade of August (2019, 2020) (Fig. 3).
Immature M. religiosa (nymph of last instars) were recorded in Lithuania for the fi rst time in 2017. In total there were 15 reports of immature M. religiosa from seven districts of southern, eastern and central parts of Lithuania (Fig. 4). Immatures were recorded earlier than adults, with the earliest reported on the 4 th (2019, Žuvintas) and 15 th of July (2019, Vilnius) and the latest on the 16 th and 12 th of September (2017, Druskininkai and Vilnius). The highest numbers of nymphs were recorded in the middle of August. Of the nymphs recorded 54% were found in natural and semi-natural habitats (heather, quarries, meadows) and 46% in anthropogenic habitats (homesteads surrounded by a green area or forest).
The locations at which M. religiosa were mainly recorded or collected were city streets and walls of buildings where they were possibly attracted to lights, suburban courtyards, and gardens. Some specimens were collected during the trapping of moths using light traps. There are fi ve records of oothecas attached to the walls of buildings and in greenhouses on fl owers, with females of M. religiosa observed close to the oothecas. There are also reports of females and males mating.
All the adults and immature M. religiosa recorded in natural environments by the authors were for dry habitats: dry sandy grasslands, heaths, bushy grassland habitats. The regular fi ndings of adult and immature M. religiosa in south and south-eastern Lithuania in natural habitats indicate the establishment of local populations of the European mantis.

Molecular analysis
We obtained COI (643 bp.) and COII (716 bp.) sequences from sixteen individuals of M. religiosa collected in Lithuania in 2016-2019 and specimens collected from The Czech Republic, Belarus and France. Four COI and 3 COII haplotypes were recorded in Lithuania. The same COI and COII haplotypes were detected in different years of the study indicating that the origin of M. religiosa in different years was the same. Sequences for M. religiosa from Belarus differed by 0-0.3% (COI) and 0-0.1% (COII) from those detected in Lithuania and that of material collected in France and The Czech Republic differed by 1.1-2.2% (COI) and 2.5-2.8 (COII). The maximum likelihood ph ylogenetic trees revealed three (COII) and three (COI) clades (Fig. 5). All specimens collected in Lithuania were in clade A together with a specimen collected in Belarus (M19) and sequences deposited in GenBank from Ukraine (Fig. 5). Sequences of M. religiosa collected in France and The Czech Republic clustered in clade B together with sequences from the GenBank obtained from specimens collected in Italy, Austria, Hungary and Slovakia (Fig. 5). The third clade C in the phylogenetic tree included both COI and COII sequences from M. religiosa collected in Germany and France.
Similarly, the January (coldest month of the year) mean air temperatures have increased in Lithuania since the year 1988 ( Fig. 7) thus creating suitable conditions for M. religiosa to overwinter.

Recent changes in climate, land use and increasing mobility have resulted in range shifts of insects in
Targeted questionnaires were used to study the distribution of M. religiosa in Lithuania, a method that was successfully used in Poland and Latvia (Liana, 2007;Pupiņš et al., 2012;Zielinski et al., 2018). A similar method was used to collect information on the distribution of the invasive mantids Hierodula patellifer a Serville, 1839 in France (Moulin, 2020) and H. tenuidentata Saussure, 1869 and Sphodromantis viridis (Forskål, 1775) in Italy (Battiston et al., 2018(Battiston et al., , 2019. Surveys based on questionnaires gather a lot of information about the spread of a target species from all over a country, but do not confi rm the presence of vital subpopulations and lack information on habitats. The spread of M. religiosa in Lithuania began in 2008 (Rimšaitė et al., 2017), when fi ve adult females were found in different parts of Vilnius for the fi rst time. The localities where most of these insects were fi rst recorded (2008)(2009)(2010)(2011)(2012)(2013)(2014)(2015) in Lithuania were near highways and transport cor-ridors and only three specimens were recorded at the edges of pine forests or peat bogs (Rimšaitė et al., 2017). This indicates that M. religiosa may be transported on vehicles along transport corridors. During the fi rst period of this study (2008)(2009)(2010)(2011)(2012)(2013)(2014)(2015) the highest numbers of records were for the surroundings of Vilnius and southern part of the country (Rimšaitė et al., 2017). The European mantis spread during the period 2015-2020 from south, south-east and east Lithuania to south-west and west Lithuania. The largest numbers of reports were for the biggest towns, which can be explained by the specifi city of the data collection, due to the large number of respondents in metropolitan areas and not to the fact that they are population hotspots.
Mantis religiosa inhabited regions in south-eastern Poland until 2007(Liana, 2007 and recently sprea d northwards in this country (Durak et al., 2018;Kadej et al ., 2018;Zieliński et al., 2018), as is reported in Germany and The Czech Republic (Landeck et al., 2013;Linn & Gr iebelen, 2015;Mückstein, 2016). One individual M. religiosa was recorded for the fi rst time in Latvia at Lielvārde in 2008, th e same yea r as in Lithuania and several individuals were also found in 2010 (Pupiņš et al., 2012). This species reached Estonia ten years later, in 2019 (27 th of August), the  Table S1.
fi rst adult female was recorded close to Pärnu (58.27831, 24.579328) (Truuverk, 2019) and this is now the northernmost record for M. religiosa. Before this the northernmost record for M. religiosa was in Russia at Severnoje in the Kostroma distr. (58.03002, 41.42999) (Bolshakov et al., 2010). In Bel arus and Russia, the spread of this species to the north also started in 2008 (Kulak, 20 09;Aleksandrowicz, 2011;Ostrovsky, 2017;Serzhantova et al., 2017Serzhantova et al., , 2019. Development time and moulting periods of M. religiosa differ in different countries because the temperatures there also differ (Liana, 2007;Linn & Griebeler, 2016). Adult M. religiosa emerge in May in warm Mediterranean regions, whereas nymph development is typically completed in late July or August in colder countries such as Poland and Germany (Liana, 2007;Linn & Griebeler, 2016). The same periodicity in the appearance of imagines was recorded in Lithuania, with the fi rst adults recorded in the fi rst decade of August and only one record of an adult mantid at the end of July in 2020, while juveniles were observed from the fi rst decade of July. The phenology of the life cycle of the perennial and viable populations in Germany and Poland confi rms that it is likely this species will adapt to local conditions in Lithuania.
Habitats of M. religiosa in Central Europe are similar to those of other xerophilous or xerothermic insects, such as bush-crickets Phaneroptera falcata (Liana, 2007;Bolshakov et al., 2010;Landeck et al., 20 13). Orthoptera usually form an important part of the adult diet of M. religiosa (Landeck et al., 2013;Breitenmo sen, 2016). Bush-crickets were recorded in south Lithuania in 2008 for the fi rst time (Ivinskis & Rimšaitė, 2008b), the same year as M. religiosa. Now bush-crickets are distributed throughout Lithuania and were recorded in south-eastern Latvia in 2011, 300 km north-east of the locality of the fi rst record in Lithuania. This indicates that the speed of spread of this species can be approximately 100 km per year (Budrys et al., 2015), although other authors indicate a much lower speed (Kočárek et al., 2008).
Typical localities of M. religiosa in Poland are clearings coniferous forests or grasslands at the edges of these forests, where the microclimate is warmer (Liana, 2007). Due to climate change and increase in temperature the role of such habitats become less important as it is currently colonizing new suitable habitats − waste and fallow lands (Liana, 2007). Studies on the hab itat requirements of M. religiosa in Germany and Italy indicate the importance of microhabitats: females prefer solid substrates with high heat-storing capacities for egg deposition, as it results in faster egg development (Batisttiston & Fontana, 2010;Li nn & Griebeler, 2016). As M. religiosa locates more suitable habitats, local populations will become established and as a consequence the species will spread further northwards. Across Europe air temperatures have been increasing since the 20 th century, the European average temperatures indicate that 7 of the top 10 warmest years were recorded in the period starting in the year 2000 and there is a clear upward trend in annual average temperatures over the last few decades (van der Schrier et al., 2013). In this period, winters and springs were warmer (~ 1.7-2.0°C), whereas summer and autumn temperatures changed less. The last decade of the 20 th century was distinguished by a unique climatic phenomenon of several successive warm winters (1988/1989-1994/1995). For the last two hundred years, eastern Europe has never experienced such a long series of anomalously warm winters (Bukantis, 2001). This creates the preconditions for the establishment of overwintering populations of southern species of insects in this region.
We found adult M. religiosa of both sexes and since 2017 also juveniles, which indicates this species can successfully overwinter in Lithuania. The presence for several years of juveniles and adults in the same natural areas in south and south-eastern Lithuania indicate that there are local population of M. religiosa in Lithuania and this species is now established here and continues to spread.
As in its distribution elsewhere there are two colour forms of M. religiosa in Lithuania: grass-green and brown shading from yellow to dark brown. The different colours are possibly related to the places where they can be wellhidden and indirectly related to climatic conditions and colour of the substrate (Battiston & Fontana, 2010), but no t to sex (Lopez, 1998), which is in accordan ce with our results. Our data revealed that females dominate both colour forms of the European mantis.
Genetic diversity, based on mtDNA, of the M. religiosa in Lithuania is poor with only few haplotypes, which indicates they all originated from the same locality. Possible routes followed by M. religiosa in its northward spread in Europe are reported by Linn & Griebelen (2015). They anal ysed four mtDNA genes of European mantis throughout Europe, except for Northern Europe and the Baltic countries, and detected three main clades of M. religiosa in Europe. The fi rst clade (A), called the East lineage, includes those from the Ukraine the sequences of which are similar to those of the Lithuania and Belarus specimens in the present study. This indicates that M. religiosa spread from Ukraine and Belarus to Lithuania. The second clade (B) determined by Linn & Griebelen (2015) includes all central European, Italian and east German haplotypes, hereafter called the Central lineage (in the present study it includes sequences from mantids collected in France and The Czech Republic). The third clade (C) includes the French and west German haplotypes, hereafter called West lineage. This clustering refl ects genetic isolation by distance during glacial periods (Hewitt, 1999) and the major ice ag e refugia (Linn & Griebelen, 2015). As presum ed, M. religiosa spread to Lithuania from one direction, the southeast (Ukraine). In contrast, the German populations of the European mantis originate from two directions: eastern France (West lineage) and central Europe (Central lineage).
The haplotype diversity reported in populations of M. religiosa is very low and possibly the result of colonization by a small number of individuals (Taberlet et al., 1998;Hewitt, 199 9;Dlugosch & Parker, 2008) as in the case of Lithuania.
Climate change is one of the reasons for the current northward spread of M. religiosa and other species from southern Europe, but not the only one. The distribution of M. religiosa in central Europe was also limited by environmental conditions (Liana, 2007) and the spread of this species has been possible because of new types of suitable environment. Further research on M. religiosa in Lithuania is needed to clarify which habitat features are favourable for these insects and which microhabitats it colonizes fi rst.