Introduction
Bumblebees are very important pollinators of both crops and native plants and consist of 250 species worldwide (Michener, 2000; Williams, 1998). After the improvements in mass rearing methods, bumblebees are increasingly used for pollination of crops, primarily for tomato pollination in greenhouses (Velthuis & van Doorn, 2006). It is estimated that more than two million commercially reared bumblebee colonies are used for the pollination globally each year (Lecocq, Coppee, Michez, Brasero , Rasplus, Valterova, & Rasmont, 2016). Currently, five species of bumblebees are reared commercially. The main species is the buff-tailed bumblebee, Bombus terrestris L. It is naturally distributed mainly in Europe, in coastal North Africa, and in West and Central Asia. Since the late 1980s, commercially rearedB. terrestris colonies have been transported to many countries, in every continent except Australia and North America (Rasmont, Coppee, Michez, & Meullemeester, 2008; Velthuis & van Doorn, 2006). Shortly after commercial introduction widely beyond its natural area of distribution, it was recognized that B. terrestris is invasive and may disturb local ecosystems including; competition for nest sites and food resources with local bee fauna, genetic contamination of localBombus spp ., and spread of parasites and pathogens (Dafni, Kevan, Gross, & Goka, 2010).
B. terrestris comprises nine recognized subspecies which differ in morphology (particularly the pattern of coat colour), genetic, behavior, colony size, etc. and inhabit separate geographic regions, sometimes overlapping. Although many subspecies of B. terrestriswere used in the early years of commercial rearing, today Bombus terrestris dalmatinus has become the main commercialized subspecies due to the suitable characteristics for mass rearing such as the large size of workers and high success rate of colonies. Bombus terrestris terrestris is also currently produced by some companies. Commercial produced colonies probably originate from stocks collected from the Turkey, Greece and Balkan Peninsula (B. t. dalmatinus ) and the northern Europe (B. t. terrestris ) (Lecocq, Rasmont, Harpke, & Schweiger, 2016; Rasmont, Coppee, Michez, & Meullemeester, 2008; Velthuis & van Doorn, 2006).
There are many characteristics which may contribute to making B. terrestris invasive including; high migration ability, early seasonal emergence, high adaptability under adverse climatic conditions in various habitats, generalist or polylectic foraging strategies, thermoregulatory metabolism that allows it to be active at low temperatures, high reproductive abilities, being able to regulate life cycle in a year with or without hibernation or aestivation (Dafni, Kevan, Gross, & Goka, 2010). Additionally, commercial B. terrestris colonies produce more gynes (queens) and are better competitors than the local conspecific populations. (Gosterit & Gurel, 2005; Ings, Ward, & Chittka, 2006). A single commercial B. terrestris colony is capable of producing more than a hundred gynes (new queens) and males which potentially could escape from greenhouses. Therefore, commercially produced B. terrestris populations are able to establish and spread into the wild of both native and non-native regions. In fact, commercial greenhouse B. terrestris populations have become established outside its native range in many countries such as, New Zealand, Tasmania, Japan, Chile and Argentina (Buttermore, 1997; Goka, 2010; Montalva, Dudley, Arroyo, Retamales, & Abrahamovich, 2011; Morales, Arbetman, Cameron, & Aizen, 2013). Within its native range, the large-scale use of B. terrestris in greenhouse also raises concerns about the genetic introgression between commercial and nativeB. terrestris populations which include recognized subspecies or distinct populations. For instance, Kraus, Szentgyorgyi, Rozej, Rhode, Moron, Woyciechowski, & Moritz, (2011) in Poland found strong genetic introgression from the sampled greenhouse populations into the adjacent populations also found that more distant populations were much less affected by genetic introgression from the greenhouses. Goulson (2010) suggested that since the beginning of the bumblebee trade in the 1980s,Bombus terrestris audax which is endemic to Britain and Ireland may have already merged with commercial produced B. t. dalmatinusand B. t. terrestris through introgression resulting in a single population. Recently potential hybrids between managed B. t. terrestris /dalmatinus and Bombus terrestris lusitanicus (endemic in Iberian Peninsula) have been reported by Cejas, Ornosa, Munoz, & De la Rua, (2018) and Seabra et al. (2019).
B. t. dalmatinus is the only native B. terrestrissubspecies that is found in Turkey. This subspecies of bumble bees are the most widespread and live at different altitudes and habitats in the country. Therefore, there are probably several ecotypes of this subspecies such as aestivated and hibernated populations, each adapted to specific ecological conditions; from the Mediterranean coastal areas to the high mountain conditions of the central Anatolian region. (Gurel, Gosterit, & Eren, 2008; Ozbek, 1997; Yeninar, Duchateau, Kaftanoglu, &Velthuis, 2000). On the other hand, commercially produced B. terrestris colonies began to use as a tomatoes pollinator in 1997. The number of commercial colonies used has increased enormously year by year and annually reached to almost 300 000 colonies in 2018 in Turkey. Because of the lack of regulations with respect to introduction of non - native B. terrestris subspecies, the movement of commercial B. terrestris colonies has largely been without risk assessment and poses a significant disease transmission and hybridization risk. The consequences of commercialization on wild populations, the extent of their spread, the degree of genetic introgression into wild populations are not well understood. Therefore, in this study we sampled B. terrestris from greenhouses (belonging to all local and global companies), within the 5 km areas surrounding the greenhouses and more distant native populations in the Mediterranean region to determine the genetic structure of native and commercial B. terrestris populations and to better understand genetic introgression of commercial greenhouse B. terrestrispopulations into the native B. t. dalmatinus populations using twenty microsatellite markers and two mitochondrial genes (COIand cyt b ).