© 2008 The Authors Journal compilation © 2008 The Royal Entomological Society 113 Insect Molecular Biology (2008) 17(2), 113–124 doi: 10.1111/j.1365-2583.2007.00787.x Blackwell Publishing Ltd Multiple invasions of Errantivirus in the genus Drosophila A. Ludwig*, V. L. da S. Valente* and E. L. S. Loreto*† *Programa de Pós-Graduação em Genética e Biologia Molecular, Departamento de Genética, Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, Rio Grande do Sul, Brazil; and †Departamento de Biologia, Universidade Federal de Santa Maria (UFSM), Campus Universitário, Santa Maria, Rio Grande do Sul, Brazil Abstract Aiming to contribute to the knowledge of the evolutionary history of Errantivirus, a phylogenetic analysis of the env gene sequences of Errantivirus gypsy, gtwin, gypsy2, gypsy3, gypsy4 and gypsy6 was carried out in 33 Drosophilidae species. Most sequences were obtained from in silico searches in the Drosophila genomes. The complex evolutionary pattern reported by other authors for the gypsy retroelement was also observed in the present study, including vertical transmission, ancestral polymorphism, stochastic loss and horizontal transfer. Moreover, the elements gypsy2, gypsy3, gypsy4 and gypsy6 were shown to have followed an evolutionary model that is similar to gypsy. Fifteen new possible cases of horizontal transfer were suggested. The infec- tious potential of these elements may help elucidate the evolutionary scenario described in the present study. Keywords: horizontal transfer, gypsy, gypsy2, gypsy3, gypsy4, gypsy6, gtwin, complex evolution, transposable elements, env gene. Introduction The errantiviruses are insect long terminal repeat (LTR) retrotransposons, characterized by the presence of the env gene, in addition to the gag and pol genes. These elements show considerable structural similarity to vertebrate retro- viruses and are named insect endogenous retroviruses, which belong to the Metaviridae family, according to the International Committee on Taxonomy of Viruses (ICTV) (Boeke et al., 1999). The most commonly studied errantivirus is the gypsy retroelement of Drosophila melanogaster, which shows infectious properties under specific conditions (Kim et al., 1994; Song et al ., 1994). Sequences homologous to the gypsy retroelement are widely distributed across the genus Drosophila (Stacey et al ., 1986; Terzian et al ., 2000; Vázquez- Manrique et al., 2000; Herédia et al., 2004, 2007) and related elements have been identified in the D. melanogaster genome sequence (Jurka, 2000; Bowen & McDonald, 2001; Kaminker et al., 2002; Kapitonov & Jurka, 2003). Like any typical endogenous retrovirus, gypsy is vertically transmitted as part of the host genome, although horizontal transfer (HT) events of this retroelement have been suggested for several Drosophila species (Alberola & de Frutos, 1996; Terzian et al., 2000; Vázquez-Manrique et al., 2000; Herédia et al., 2004). It has been hypothesized that this ability of the gypsy element to undergo HT is related to the presence of the functional env gene, which in turn is responsible for the infectious properties of retroviruses (Mejlumian et al., 2002; Herédia et al., 2004). This study analyses an approximately 500-bp region of the env gene of the gypsy and related retroelements in several Drosophilidae species. New sequences for this region were obtained by cloning and sequencing of the gypsy element in the D. flavopilosa group species and by in silico searches of sequences homologous to gypsy, gypsy2, gypsy3, gypsy4 and gypsy6 from the Drosophila genome sequences currently available. The sequences obtained were analysed together with those deposited in GENBANK. Twenty-seven probable HT cases were identified, of which 12 have previously been described (Alberola & de Frutos, 1996; Vázquez-Manrique et al., 2000; Herédia et al., 2004; Ludwig & Loreto, 2007). The results of the present study agree with the findings by Herédia et al. (2004), which presented a complex evolutionary model for gypsy. It was observed that gypsy2, gypsy4 and gypsy6 seem to be involved in HT events. Results gypsy in the D. flavopilosa group The D. flavopilosa group is formed by species that present a considerably restricted ecology. These species use Received 24 August 2007; accepted after revision 3 December 2007; first published online 15 February 2008. Correspondence: Elgion Lúcio Silva Loreto, Departamento de Biologia, Universidade Federal de Santa Maria (UFSM), Campus Universitário, Santa Maria, Rio Grande do Sul, Brazil, 97105-900. Tel.: +55 55 3220 8912; fax: +55 55 3220 8912; e-mail: elgionl@gmail.com