Identification key for drosophilid species (Diptera, Drosophilidae) exotic to the Neotropical Region and occurring in Brazil

Thirteen species of drosophilid exotic to the Neotropical Region are recorded in Brazil, and some of them are highly invasive and threaten significantly fruit cultures. We provide an illustrated key for identifying these species, and briefly discuss their taxonomic status, distribution, and occurrence in the Neotropics. The key should not only support newcomers to the study of Drosophila but also facilitate their identification by those interested in insects associated with cultivated areas. A R T I C L E I N F O Article history: Received 20 October 2019 Accepted 07 December 2019 Available online 31 January 2020 Associate Editor: Claudio Carvalho


Introduction
The Taxonomic Catalog of the Brazilian Fauna (TCBF) records 305 drosophilid species in this country . Most are neotropical and ecologically restricted to a particular type of vegetation. Thirteen of these species, however, are exotic to the Neotropical Region and widely distributed in the world. Some (e.g., Drosophila melanogaster Meigen and D. simulans Sturtevant) possibly arrived in Brazil in the 16th century, transported by slave ships from Africa. Others reached the country later, throughout trade ships and airplanes. From the late 20th century, four new arrivals in the Neotropics were accurately recorded in the earlier stages of invasion: D. malerkotliana Parshad and Paika , Zaprionus indianus Gupta (Vilela, 1999), D. nasuta Lamb (Vilela and Goñi, 2015) and D. suzukii Matsumura (Deprá et al., 2014).
The impacts of biological invasions have been widely recognized since the seminal book The ecology of invasions by animals and plants (Elton, 1958). Ecological interactions between invasive and native species, like predation and competition, often affect the population dynamics (births, deaths, migration) of native species and bring severe negative consequences for biodiversity (Millennium Ecosystem Assessment, 2005). Moreover, invasive species can also cause economic troubles. In Brazil, annual losses to major crops caused by alien species are estimated to about US$1.6 billion (ca. 16% caused by dipterans), and for USA and India this value is even higher (Oliveira et al., 2013). Therefore, it is crucial to understand the biology and distribution of exotic species (Sakai et al., 2001).
Taxonomic identification based on morphological characters is an effective way of determining many drosophilid species, especially the non-natives in a particular region. However, there is a lack of identification resources that can be easily used by non-specialists. The primary dichotomous key for drosophilids in Brazil (Freire-Maia and Pavan, 1949) attended several generations of researchers, and certainly contributed for spreading the study of these flies in the country. However, it does not include the recently introduced species and needs to be taxonomically upgraded (e.g. Drosophila mirim was synonymized as Scaptodrosophila latifasciaeformis). Here, we provide an illustrated identification key for 13 exotic drosophilid species recorded in Brazil and briefly discuss their taxonomic status, distribution, and occurrence in the Neotropics.

Material and Methods
We analyzed specimens of 11 among the 13 non-neotropical drosophilids occurring in Brazil and collected data of Drosophila virilis and Scaptodrosophila lebanonensis from the literature ( Table 1). The illustrations were hand-made utilizing a camera lucida coupled to a Leica MZ75 stereoscope and followed traditional China ink scientific illustration methods reassembling those from Bächli et al. (2004). The morphological terms used in the dichotomous key are also based mainly on Bächli et al. (op cit.).
The dichotomous key does not consider the degree of phylogenetic correlation, but rather the difficulty of identifying each specimen. Thus, the species with the most striking morphological characteristics (Z. indianus with their bright stripes and D. busckii with their mesonotum patterns) were placed at the first steps of the key. Those specimens that required the rigorous analysis of bristles or sex combs (Scaptodrosophila genus and melanogaster species group) were left for the end section of the key.
The characters addressed here to identify females of D. melanogaster and D. simulans are useful for lab routine or identifying flies in the field, but insufficient for a taxonomic study because the black pigmentation on each tergite is highly variable according to genotype and grow temperature (Moreteau et al., 1995). For more accurate techniques such as egg-shape inspection or wing and thorax size ratio see Moreteau et al. (op cit.).

Discussion
Potentially colonizer drosophilids had attracted the attention of researchers for over five decades (Dobzhansky, 1965;Lewontin, 1965;David and Tsacas, 1981;Parsons, 1983), and it is well established that these flies can be used as models in studies of biological invasions (Gibert et al., 2016). The species registered here are widely distributed not only in South America but also in other regions of the world (Bächli, 2019). The section below provides information about the origin (David and Tsacas, 1981), distribution (Brake and Bächli, 2008;Bächli, 2019), and general characteristics of each species.

Genus Drosophila Fállen
The genus Drosophila includes approximately half of the 4,000 species in the family Drosophilidae. Its members have adapted to and radiated in a variety of niches (Markow and O'Grady, 2008), and currently divided into nine subgenera (O'Grady and DeSalle, 2018).

Subgenus Dorsilopha Sturtevant
This subgenus of oriental origin (Toda, 1986) consists of only four species, including the cosmopolitan Drosophila busckii. While many common drosophilids prefer decaying fruit, D. busckii larvae seem to have a broader niche (Atkinson and Shorrocks, 1977) and were recorded feeding on many different vegetables (Valadão et al., 2019), animal excrement and meat (Sturtevant, 1921). This species is very abundant in temperate places, where its competitive ability is apparently higher (David and Tsacas, 1981). In Brazil, D. busckii is rare in natural habitats but can be found in disturbed areas .

Subgenus Drosophila Fállen
This is the largest subgenus of Drosophila, supporting about 80% of its ca. 2000 species. Therefore, this subgenus is organized in 47 species groups, which are an informal taxonomic rank (Sturtevant, 1942) that organize the diversity within large clades of drosophilids.
The immigrans species group, probably of oriental origin, currently contains about 100 nominal species and two of them are recorded in Brazil. The cosmopolitan Drosophila immigrans is abundant in the Palearctic and Nearctic Regions (David and Tsacas, 1981) and has been collected in Brazilian natural and modified environments since the pioneer studies of Pavan (1959). In this country, the abundance of D. immigrans seems to be low at warmer areas  but increases in the colder southern regions (Hochmüller et al., 2010). Drosophila nasuta, on the other hand, is widespread and abundant in the tropical parts of Africa (David and Tsacas, op cit.) and has been recently recorded in the Neotropics (Vilela and Goñi, 2015). In forest patches located in the Brazilian savanna, where D. immigrans remains rare (data not shown), D. nasuta can reach 20% relative abundance of drosophilids during Summer months (Leão et al., 2017). These findings support the hypothesis raised by David and Tsacas (op cit.) that D. immigrans is cold-adapted, whereas D. nasuta is a tropical species.
The virilis species group consists of about thirteen described species that are typically found in a boreal distribution (Markow and O'Grady, 2006). The group appears to have originated in Asia as a species of the temperate deciduous forest associated predominantly with riparian communities (Throckmorton, 1975). Only Drosophila virilis Sturtevant has been recorded worldwide in wine production areas as well as in breweries (Bächli et al., 2004).

Subgenus Sophophora Sturtevant
This subgenus currently contains 344 species organized in nine species groups. The Drosophila melanogaster species group, the largest of this subgenus, contains almost 200 species widely distributed in the Oriental Region and adjacent areas, but some lineages reached the African continent and radiated. Six species of this group are recorded in Brazil.
The sibling African species Drosophila melanogaster and D. simulans probably are the most studied examples of widespread drosophilids (Capy et al., 2004). D. melanogaster first colonized Eurasia (10,000-15,000 years ago) and later spread to America and Australia. It is generally admitted that this human commensal species travel easily with fruit shipments, mostly as larvae or pupae on rotting material. D. simulans expanded worldwide more recently, probably a few centuries ago, establishing in natural and disturbed environments throughout the invaded areas. In Brazil, both species arrived probably as a result of the slave trade. Currently, D. simulans is a widespread species in all sorts of environments, whereas D. melanogaster is rare in natural habitats . Drosophila ananassae, probably native of the Orient, is a peridomestic species occurring in tropical and subtropical areas of the world. In Brazil, it is rare in natural environments but can be found in open types of vegetation . We have been collecting this species in urban orchards, near Brasília.
The oriental species D. kikkawai supports a cosmopolitan status (Pinheiro and Valente, 1993), but it is not clear when it was introduced in South America and Africa and nor if this process was mediated by humans (Ashburner, 1989). In Brazil, this species is found in both natural  and urban environments (Costa et al., 2003), usually in low abundances (but see Cavasini et al., 2014).
Drosophila malerkotliana, also native of the Orient, was described from the Malerkotla region in India by Parshad and Paika, in 1964. Besides being widespread in the Orient, it also occurs in Africa and Americas, where it was introduced in recent decades Sene et al. 1980;David and Tsacas 1981;Castrezana et al., 2010). In Brazil, it occurs in both natural environments (Medeiros and Klaczko, 2004;Torres and Madi-Ravazzi, 2006;Tidon, 2006;Schmitz et al., 2010), and urbanized sites .
Finally, a species that is currently meriting the status of cosmopolitan is D. suzukii, known as the spotted-wing Drosophila due to the dark spot on the male wings. Probably native of East Asia, this species has spread to western regions: it was found in Hawaii in the early 1980s and more recently in North America and Europe, where it has reached invasive pest status (Asplen et al., 2015). The female has a long and narrow ovipositor used to infest soft-skinned fruit crops and causing enormous economic damage on commercial plantations; hence, the spotted-wing Drosophila has become an agricultural pest in several countries worldwide (Walsh et al., 2011). In Brazil, this cold-adapted species was first detected in the southern region in February 2013 (Deprá et al., 2014). As conjectured by Vilela and Mori (2014), who recorded this species in Southeast Brazil, D. suzukii is probably expanding its territory to other South American areas through the trade of cultivated soft skin fruits and the use of small wild fruits as breeding sites. Although Southern Brazil is the most climatically favorable area for D. suzukii development (Benito et al., 2016), it has already been found in natural vegetations of the tropical Brazilian Savanna, at low abundances (Leão et al., 2017).

Genus Scaptodrosophila Duda
The genus Scaptodrosophila, initially described as a subgenus of Drosophila, was formally elevated to generic rank by Grimaldi (1990). Probably originated in tropical Asia, Scaptodrosophila currently includes almost 300 species distributed in Asia, New Guinea, Australia, and Africa, with few species in the Americas and Europe. Two species of this genus have reached the Neotropical Region. Scaptodrosophila latifasciaeformis is a widespread species in tropical and subtropical areas of the world. In Brazil, it was described by Dobzhansky and Pavan (1943) as Drosophila mirim and has been collected in all types of vegetations , including mangroves in the southern region of the country . S. lebanonensis, later described as Drosophila galloi by Lourenço and Mourão (1992) and synonymized by Bächli et al. (2005), was recorded in southern Neotropical region in 1960 and we have no notice of other records in the Neotropical region after then.

Genus Zaprionus
Zaprionus is an Afrotropical genus that extended its distribution to the Australian, Oriental and Palearctic regions. Zaprionus indianus constitutes one of the most successful colonizing species of this genus (Chassagnard and Tsacas, 1993), probably because of its broad niche-width characteristics: it utilizes diverse food resources and displays adaptation to variable climatic conditions (Parkash and Yadav, 1993). The first record of Z. indianus in Brazil was in São Paulo State (Vilela, 1999) and, after that, the species was found in many Brazilian regions (Galego and Carareto, 2010). In natural areas where environmental conditions are similar to those observed in its original area in Africa, Z. indianus dominates drosophilid assemblages during the wet season (Tidon et al., 2003). It also predominates in fig plantations: among 125,00 drosophilids captured in São Paulo State, 83,339 were identified as Z. indianus . This species is highly adaptable (Mata et al., 2010) and deserves to be monitored.

Final Remarks
Most drosophilids approached in this study are widely distributed thought natural populations in many Brazilian biomes, and two species in particular, Drosophila suzukii and Zaprionus indianus, can cause great impact on cultivated areas. The identification key provided here should not only support newcomers to the study of Drosophila but also the professional that will be dealing with the elimination of the pests affecting plantation, who generally will not be Drosophila experts.