Horizontal transfer and the widespread presence of Galileo transposons in Drosophilidae (Insecta: Diptera).

Galileo is a transposon notoriously involved with inversions in Drosophila buzzatii by ectopic recombination. Although widespread in Drosophila, little is known about this transposon in other lineages of Drosophilidae. Here, the abundance of the canonical Galileo and its evolutionary history in Drosophilidae genomes was estimated and reconstructed across genera within its two subfamilies. Sequences of this transposon were masked in these genomes and their transposase sequences were recovered using BLASTn. Phylogenetic analyses were employed to reconstruct their evolutionary history and compare it to that of host genomes. Galileo was found in nearly all 163 species, however, only 37 harbored nearly complete transposase sequences. In the remaining, Galileo was found highly fragmented. Copies from related species were clustered, however horizontal transfer events were detected between the melanogaster and montium groups of Drosophila, and between the latter and the Lordiphosa genus. The similarity of sequences found in the virilis and willistoni groups of Drosophila was found to be a consequence of lineage sorting. Therefore, the evolution of Galileo is primarily marked by vertical transmission and long-term inactivation, mainly through the deletion of open reading frames. The latter has the potential to lead copies of this transposon to become miniature inverted-repeat transposable elements.

Patterns of genome size evolution versus fraction of repetitive elements in statu nascendi species: the case of the willistoni subgroup of Drosophila (Diptera, Drosophilidae).

Genome size evolution is known to be related with transposable elements, yet such relation in incipient species remains poorly understood. For decades, the willistoni subgroup of Drosophila has been a model for evolutionary studies because of the different evolutionary stages and degrees of reproductive isolation its species present. Our main question here was how speciation influences genome size evolution and the fraction of repetitive elements, with a focus on transposable elements. We quantitatively compared the mobilome of four species and two subspecies belonging to this subgroup with their genome size, and performed comparative phylogenetic analyses. Our results showed that genome size and the fraction of repetitive elements evolved according to the evolutionary history of these species, but the content of transposable elements showed some discrepancies. Signals of recent transposition events were detected for different superfamilies. Their low genomic GC content suggests that in these species transposable element mobilization might be facilitated by relaxed natural selection. Additionally, a possible role of the superfamily DNA/TcMar-Tigger in the expansion of these genomes was also detected. We hypothesize that the undergoing process of speciation could be promoting the observed increase in the fraction of repetitive elements and, consequently, genome size.

Structural and sequence diversity of the transposon Galileo in the Drosophila willistoni genome.

BACKGROUND: Galileo is one of three members of the P superfamily of DNA transposons. It was originally discovered in Drosophila buzzatii, in which three segregating chromosomal inversions were shown to have been generated by ectopic recombination between Galileo copies. Subsequently, Galileo was identified in six of 12 sequenced Drosophila genomes, indicating its widespread distribution within this genus. Galileo is strikingly abundant in Drosophila willistoni, a neotropical species that is highly polymorphic for chromosomal inversions, suggesting a role for this transposon in the evolution of its genome. RESULTS: We carried out a detailed characterization of all Galileo copies present in the D. willistoni genome. A total of 191 copies, including 133 with two terminal inverted repeats (TIRs), were classified according to structure in six groups. The TIRs exhibited remarkable variation in their length and structure compared to the most complete copy. Three copies showed extended TIRs due to internal tandem repeats, the insertion of other transposable elements (TEs), or the incorporation of non-TIR sequences into the TIRs. Phylogenetic analyses of the transposase (TPase)-encoding and TIR segments yielded two divergent clades, which we termed Galileo subfamilies V and W. Target-site duplications (TSDs) in D. willistoni Galileo copies were 7- or 8-bp in length, with the consensus sequence GTATTAC. Analysis of the region around the TSDs revealed a target site motif (TSM) with a 15-bp palindrome that may give rise to a stem-loop secondary structure. CONCLUSIONS: There is a remarkable abundance and diversity of Galileo copies in the D. willistoni genome, although no functional copies were found. The TIRs in particular have a dynamic structure and extend in different ways, but their ends (required for transposition) are more conserved than the rest of the element. The D. willistoni genome harbors two Galileo subfamilies (V and W) that diverged ~9 million years ago and may have descended from an ancestral element in the genome. Galileo shows a significant insertion preference for a 15-bp palindromic TSM.

Hosimary: a new hAT transposon group involved in horizontal transfer.

A PCR screening approach was used to search for sequences homologous to a previously described hAT transposon found in Drosophila simulans and Drosophila sechellia, named here as hosimary. In this study, 52 Drosophilidae species were analyzed and these sequences seem to be restricted to some species of the melanogaster group and Zaprionus indianus. These species present variable number of copies and most of those appear to be putatively encoding. The high hosimary sequences similarity among different species and the patchy distribution presented by this transposon strongly support the hypothesis that hosimary was horizontally transferred between the melanogaster group species and Z. indianus.

Phylogenetic relationships and macro-evolutionary patterns within the Drosophila tripunctata "radiation" (Diptera: Drosophilidae).

Despite previous efforts, the evolutionary history of the immigrans-tripunctata clade remains obscure in part due to its hypothesized origin through a rapid radiation. We performed a supermatrix analysis (3,243 base pairs) coupled with richness patterns, environmental phylogenetic signal and radiation tests in order to address phylogenetic relationships and macro-evolutionary hypotheses within this complex group of species. We propose a well-supported evolutionary scenario for the immigrans-tripunctata clade species, in which the tripunctata "radiation" was monophyletic and subdivided into three main lineages: the first including D. pallidipennis (pallidipennis group) imbedded among members of the tripunctata group; the second clustering the cardini and guarani groups; and the third grouping representatives from the tripunctata, calloptera and guaramunu groups. Therefore, we hypothesize that the tripunctata group encompasses a diphyletic taxon, with one clade including the pallidipennis group and the other showing a close affinity to the calloptera and guaramunu groups. Our results also suggest that niche evolution seems to have played a central role in the evolutionary history of the tripunctata species "radiation" allowing effective dispersion and diversification in the Neotropics, possibly in a southwards direction. Although the data as a whole support the notion that this occurred through rapid and successive speciation events, the radiation hypothesis remains to be further corroborated.

Polytene chromosomal maps of 11 Drosophila species: the order of genomic scaffolds inferred from genetic and physical maps.

The sequencing of the 12 genomes of members of the genus Drosophila was taken as an opportunity to reevaluate the genetic and physical maps for 11 of the species, in part to aid in the mapping of assembled scaffolds. Here, we present an overview of the importance of cytogenetic maps to Drosophila biology and to the concepts of chromosomal evolution. Physical and genetic markers were used to anchor the genome assembly scaffolds to the polytene chromosomal maps for each species. In addition, a computational approach was used to anchor smaller scaffolds on the basis of the analysis of syntenic blocks. We present the chromosomal map data from each of the 11 sequenced non-Drosophila melanogaster species as a series of sections. Each section reviews the history of the polytene chromosome maps for each species, presents the new polytene chromosome maps, and anchors the genomic scaffolds to the cytological maps using genetic and physical markers. The mapping data agree with Muller's idea that the majority of Drosophila genes are syntenic. Despite the conservation of genes within homologous chromosome arms across species, the karyotypes of these species have changed through the fusion of chromosomal arms followed by subsequent rearrangement events.

Detection of P element transcripts in embryos of Drosophila melanogaster and D. willistoni.

The P element is one of the most thoroughly studied transposable elements (TE). Its mobilization causes the hybrid dysgenesis that was first described in Drosophila melanogaster. While studies of the P element have mainly been done in D. melanogaster, it is believed that Drosophila willistoni was the original host species of this TE and that P was transposed to the D. melanogaster genome by horizontal transfer. Our study sought to compare the transcriptional behavior of the P element in embryos of D. melanogaster, which is a recent host, with embryos of two strains of D. willistoni, a species that has contained the P element for a longer time. In both species, potential transcripts of transposase, the enzyme responsible for the TE mobilization, were detected, as were transcripts of the 66-kDa repressor, truncated and antisense sequences, which can have the ability to prevent TEs mobilization. The truncated transcripts reveal the truncated P elements present in the genome strains and whose number seems to be related to the invasion time of the genome by the TE. No qualitative differences in antisense transcripts were observed among the strains, even in the D. willistoni strain with the highest frequency of heterochromatic P elements.

Phylogeny of the Drosophila mesophragmatica group (Diptera, Drosophilidae): an example of Andean evolution.

The mesophragmatica group of Drosophila belongs to the virilis-repleta radiation of the Drosophila subgenus. This group comprises 13 Neotropical species that are endemic to the South-American continent and seem to be fundamentally Andean in their distribution. The mesophragmatica-group phylogeny has been inferred previously by other authors based on morphological, cytological, and isozyme analyses. However, the relationships within the group have not yet been completely resolved, although its monophyletic origin has already been confirmed by molecular data. This work attempts to enhance the molecular approach to the relationships among the species of the mesophragmatica group, using both nuclear and mitochondrial markers. Phylogenetic analyses were performed using fragments of the nuclear alcohol dehydrogenase (Adh; 631 bp), alpha-methyldopa (Amd; 1211 bp), dopa-decarboxylase (Ddc; 1105 bp), and hunchback (Hb; 687 bp) genes and the mitochondrial cytochrome oxidase subunit II (COII; 672 bp) gene, and included a total of 4306 bp. The sequences obtained for eight representatives of the mesophragmatica group were analyzed both individually and in combination by distance methods, maximum parsimony, and maximum likelihood. Our results support subdivision of the mesophragmatica group into three main lineages: the first is composed of D. viracochi; the second comprises a clade grouping the sibling species D. pavani and D. gaucha; and the third encompasses D. gasici, D. brncici, and D. mesophragmatica. The best supported scenario suggests that D. viracochi is an early offshoot in the mesophragmatica group, with this and other early branchings occuring in the Pliocene/Pleistocene Epochs, possibly associated with Andean glacial refuges. Also based on the phylogenies obtained, we present a genealogical view of the evolution of previously described characters within the group.

Reassignment of Drosophila willistoni Genome Scaffolds to Chromosome II Arms.

Drosophila willistoni is a geographically widespread Neotropical species. The genome of strain Gd-H4-1 from Guadeloupe Island (Caribbean) was sequenced in 2007 as part of the 12 Drosophila Genomes Project. The assembled scaffolds were joined based on conserved linkage and assigned to polytene chromosomes based on a handful of genetic and physical markers. This paucity of markers was particularly striking in the metacentric chromosome II, comprised two similarly sized arms, IIL and IIR, traditionally considered homologous to Muller elements C and B, respectively. In this paper we present the cytological mapping of 22 new gene markers to increase the number of markers mapped by in situ hybridization and to test the assignment of scaffolds to the polytene chromosome II arms. For this purpose, we generated, by polymerase chain reaction amplification, one or two gene probes from each scaffold assigned to the chromosome II arms and mapped these probes to the Gd-H4-1 strain's polytene chromosomes by nonfluorescent in situ hybridization. Our findings show that chromosome arms IIL and IIR correspond to Muller elements B and C, respectively, directly contrasting the current homology assignments in D. willistoni and constituting a major reassignment of the scaffolds to chromosome II arms.

BuT2 is a member of the third major group of hAT transposons and is involved in horizontal transfer events in the genus Drosophila.

The hAT superfamily comprises a large and diverse array of DNA transposons found in all supergroups of eukaryotes. Here we characterized the Drosophila buzzatii BuT2 element and found that it harbors a five-exon gene encoding a 643-aa putatively functional transposase. A phylogeny built with 85 hAT transposases yielded, in addition to the two major groups already described, Ac and Buster, a third one comprising 20 sequences that includes BuT2, Tip100, hAT-4_BM, and RP-hAT1. This third group is here named Tip. In addition, we studied the phylogenetic distribution and evolution of BuT2 by in silico searches and molecular approaches. Our data revealed BuT2 was, most often, vertically transmitted during the evolution of genus Drosophila being lost independently in several species. Nevertheless, we propose the occurrence of three horizontal transfer events to explain its distribution and conservation among species. Another aspect of BuT2 evolution and life cycle is the presence of short related sequences, which contain similar 5' and 3' regions, including the terminal inverted repeats. These sequences that can be considered as miniature inverted repeat transposable elements probably originated by internal deletion of complete copies and show evidences of recent mobilization.

Rhinoleucophenga joaquina sp. nov. (Diptera: Drosophilidae) from the Neotropical region.

The genus Rhinoleucophenga Hendel comprises 18 nominal species and has a Neotropical and Nearctic distribution. A new species was found in a collection in Florianópolis, SC, Brazil, and is described as Rhinoleucophenga joaquina sp. nov. The specimens found were reared from flowers of Dyckia encholirioides (Bromeliaceae), collected in a region of restinga on the dunes. The use of flowers as breeding site for Rhinoleucophenga has never been reported before.

The LTR retrotransposon micropia in the cardini group of Drosophila (Diptera: Drosophilidae): a possible case of horizontal transfer.

The presence of the micropia retroelement from the Ty1-copia family of LTR retroelements was investigated in three species of the Drosophila cardini group. Southern blot analysis suggested the existence of at least four micropia copies in the genomes of D. cardinoides, D. neocardini and D. polymorpha populations. The high sequence similarity between dhMiF2 and Dm11 clones (micropia retroelements isolated from D. hydei and D. melanogaster, respectively) with micropia sequences amplified from D. cardini group genome supports the hypothesis that this retroelement plays an active role in horizontal transfer events between D. hydei and the D. cardini group.

Changes in Brazilian Drosophilidae (Diptera) assemblages across an urbanisation gradient.

The present study aims to characterise the assemblages of Drosophilidae sampled from four sites subject to different levels of urbanisation, in the city of Florianópolis, South Brazil. Five samples were collected between August 2000 and August 2001 and five between August 2002 and August 2003 in each site. A gradual change in the dominant species was observed in the urbanisation gradient, with an increase in relative abundance of exotic species in urban areas. Among the native species, the relative abundance of Drosophila capricorni Dobzhansky & Pavan had an expressive decrease. In the other hand the richness in the low urbanisation site increased, probably due to the presence of urban gardens. The diversity index did not vary significantly among sites, but its correlation with evenness and richness changed.

Study of the Drosophilidae (Diptera) communities on Atlantic Forest islands of Santa Catarina State, Brazil.

A study of the community dynamics of Drosophilidae was carried out in six insular communities and two others on the mainland. Seasonal collections were carried out throughout two years in Santa Catarina State, southern of Brazil. The diversity index calculations show high values when compared with temperate climate communities. The sites on the mainland (Serra do Tabuleiro) presented the highest diversity, which was measured by the Diversity Index (H'). These sites are covered by primary Atlantic Forest and theoretically should have a higher variation of ecological niches. A dendogram showing the similarity between the communities, calculated by Morisita Index, points to a level of similarity equal to 60% for all communities. In this diagram, we can see two clades: one on the mainland and the other on the islands. The six island sites are grouped into one clade and separated into two subclades, one including the sites on Santa Catarina Island and the other consisting of the islands adjacent to this last and very much larger one. These groupings show the very important role of the spatial component on the prediction of the structure of the communities. This fact raises the discussion about the high complexity of the Atlantic Forest ecosystem and consequently the unpredictability of its fauna, highlighting the need of its conservation.

Inversion polymorphism and a new polytene chromosome map of Zaprionus indianus Gupta (1970) (Diptera: Drosophilidae).

Zaprionus indianus is a recent invader in Brazil and was probably introduced from the West Afrotropical zone. So far, studies regarding its chromosomal polymorphism were limited to India. We found that Brazilian populations were very different from Indian ones. Five new inversions have been discovered. In(II)A, already described in India, where it is quite common, has also been found in Brazil, where it is very rare. The X-chromosome has three inversions; In(X)Na, In(X)Ke and In(X)Eg, which are frequent in all Brazilian populations studied. In every case, we observed strong linkage disequilibrium among these gene arrangements. During the primary collection period (2001-2002), we noticed a significant positive correlation between the frequency of these inversions and latitude, but this was not confirmed in later investigations. Rearrangement In(IV)EF was also common in all populations, while inversion In(V)B was only found in southern populations. Our data suggest that the founders that recently invaded Brazil were polymorphic for the six inversions observed. The place of origin might be identified more precisely by investigating West African populations. In order to facilitate further investigations, we present an updated polytene chromosome photomap, locating the breakpoints of every inversion observed in Brazilian populations.

Taxonomic survey of Drosophilidae (Diptera) from mangrove forests of Santa Catarina Island, Southern Brazil.

Assemblages of drosophilids have been characterised in several environments of the Brazilian territory, like the Atlantic Rain Forest, urban areas, cerrados, the Amazon Forest, and others. The present survey is the first attempt to characterise the fauna of Drosophilidae in mangrove forests, an environment typical of tropical coasts worldwide. Twenty-eight samples were collected from the three main mangrove forests of Santa Catarina Island, southern Brazil, using banana-baited traps hung in trees. Samples were taken in January (summer), April (autumn), July (winter) and October (spring) between July 2002 and July 2005. In total, 82,942 specimens of drosophilids were caught, belonging to 69 species of six genera - Amiota Loew, Drosophila Fallén, Leucophenga Mik, Scaptodrosophila Duda, Zaprionus Coquillett and Zygothrica Wiedemann. The high abundance of D. simulans Sturtevant was remarkable, with some notable peaks of D. malerkotliana Parshad & Paika in autumn samples. Other common species were Zaprionus indianus Gupta, D. mediostriata Duda and D. willistoni Sturtevant. We also collected 45,826 flies of family Curtonotidae, the sister-group of Drosophilidae virtually absent in other environments. The assemblages of drosophilids were very similar in the three mangrove forests surveyed, despite the different surrounding environments. In general, the species sampled in the mangroves were the same as those observed in the surrounding environments, but in varying abundances. This suggests that drosophilids are differently affected by environmental pressures operating in mangroves.

Molecular phylogeny of the subgenus Drosophila (Diptera, Drosophilidae) with an emphasis on Neotropical species and groups: a nuclear versus mitochondrial gene approach.

The genus Drosophila has played an essential role in many biological studies during the last 100 years but much controversy and many incompletely addressed issues still remain to be elucidated regarding the phylogeny of this genus. Because information on the Neotropical species contained in the subgenus Drosophila is particularly incomplete, with this taxonomic group being underrepresented in many studies, we designed a study to answer some evolutionary questions related to these species. We subjected at least 41 Drosophilidae taxa to a phylogenetic analysis using a 516-base pair (bp) fragment of the alpha-methyldopa (Amd) nuclear gene and a 672 bp fragment of the mitochondrial cytochrome oxidase subunit II (COII) gene both individually and in combination. We found that the subgenus Drosophila is paraphyletic and subdivided into two main clusters: the first containing species traditionally placed in the virilis-repleta radiation and the second assembling species of the immigrans-Hirtodrosophila radiation. Inside the first of these clusters we could detect the monophyly of both the flavopilosa (the sister-clade of the annulimana group) and the mesophragmatica (closely related to the repleta group) species groups. Concerning the immigrans-Hirtodrosophila lineage, Zaprionus, Liodrosophila, Samoaia, and Hirtodrosophila were the early offshoots, followed by the immigrans, quinaria, testacea, and funebris species groups. The tripunctata radiation appears to be a derived clade, composed of a paraphyletic tripunctata group, intimately interposed with members of the cardini, guarani, and guaramunu species groups. Overall, the COII gene yielded a poor phylogenetic performance when compared to the Amd gene, the evolutionary hypothesis of which agreed with the total evidence tree. This phenomenon can be explained by the fast saturation of transitional substitutions in COII, due to strong biases in both base composition and substitution patterns, as also by its great among-site rate variation heterogeneity.

Rhinoleucophenga joaquina sp. nov. (Diptera: Drosophilidae) from the Neotropical region / Rhinoleucophenga joaquina sp. nov. (Diptera: Drosophilidae) da região Neotropical

O gênero Rhinoleucophenga Hendel compreende 18 espécies nominais e possui distribuição Neotropical e Neártica. Uma nova espécie foi encontrada em coleta realizada em Florianópolis, SC, e é descrita como Rhinoleucophenga joaquina sp. nov. Os espécimes encontrados emergiram de flores de Dyckia encholirioides (Bromeliaceae) coletadas em região de restinga sobre dunas. Este é o primeiro registro da utilização de flores como sítio de oviposição para Rhinoleucophenga.

The genus Rhinoleucophenga Hendel comprises 18 nominal species and has a Neotropical and Nearctic distribution. A new species was found in a collection in Florianópolis, SC, Brazil, and is described as Rhinoleucophenga joaquina sp. nov. The specimens found were reared from flowers of Dyckia encholirioides (Bromeliaceae), collected in a region of restinga on the dunes. The use of flowers as breeding site for Rhinoleucophenga has never been reported before.

Detection of P element transcripts in embryos of Drosophila melanogaster and D. willistoni

The P element is one of the most thoroughly studied transposable elements (TE). Its mobilization causes the hybrid dysgenesis that was first described in Drosophila melanogaster. While studies of the P element have mainly been done in D. melanogaster, it is believed that Drosophila willistoni was the original host species of this TE and that P was transposed to the D. melanogaster genome by horizontal transfer. Our study sought to compare the transcriptional behavior of the P element in embryos of D. melanogaster, which is a recent host, with embryos of two strains of D. willistoni, a species that has contained the P element for a longer time. In both species, potential transcripts of transposase, the enzyme responsible for the TE mobilization, were detected, as were transcripts of the 66-kDa repressor, truncated and antisense sequences, which can have the ability to prevent TEs mobilization. The truncated transcripts reveal the truncated P elements present in the genome strains and whose number seems to be related to the invasion time of the genome by the TE. No qualitative differences in antisense transcripts were observed among the strains, even in the D. willistoni strain with the highest frequency of heterochromatic P elements.

O elemento P é um dos elementos transponíveis (TE) mais amplamente estudado. Sua mobilização causa a disgenesia do híbrido que foi primeiramente descrita em D. melanogaster. Apesar dos estudos sobre o elemento P terem sido realizados principalmente com D. melanogaster, acredita-se que D. willistoni foi a espécie hospedeira original deste TE e que ele se transpôs para o genoma de D. melanogaster por transferência horizontal. Nosso estudo visou a comparação do comportamento transcripcional do elemento P em embriões de D. melanogaster, que é a hospedeira recente, com o de embriões de duas linhagens de D. willistoni, uma espécie que é, a longo tempo, hospedeira do elemento P. Em ambas as espécies foram detectados transcritos potenciais da transposase, enzima responsável pela mobilização do TE, bem como transcritos do repressor de 66-kDa e de seqüências truncadas e antisenso, os quais podem ter a habilidade de prevenir a mobilização de TEs. Os transcritos truncados refletem os elementos P truncados presentes no genoma das linhagens e cujo número parece relacionado com o tempo de invasão do genoma pelo TE. Nenhuma diferença qualitativa de transcritos antisenso foi observada entre as espécies, mesmo na linhagem de D. willistoni com alta freqüência de elemento P heterocromático.

Drosophilids (Diptera) from an Atlantic Forest Area in Santa Catarina, Southern Brazil / Drosofilídeos (Diptera) de uma Área de Floresta Atlântica em Santa Catarina, Sul do Brasil

The present work aims at knowing the faunal composition of drosophilids in forest areas of southern Brazil. Besides, estimation of species richness for this fauna is briefly discussed. The sampling were carried out in three well-preserved areas of the Atlantic Rain Forest in the State of Santa Catarina. In this study, 136,931 specimens were captured and 96.6 percent of them were identified in the specific level. The observed species richness (153 species) is the largest that has been registered in faunal inventories conducted in Brazil. Sixty-three of the captured species did not fit to the available descriptions, and we believe that most of them are non-described species. The incidence-based estimators tended to give rise to the largest richness estimates while the abundance based give rise to the smallest ones. Such estimators suggest the presence from 172.28 to 220.65 species in the studied area. Based on these values, from 69.35 to 88.81 percent of the expected species richness were sampled. We suggest that the large richness recorded in this study is a consequence of the large sampling effort, the capture method, recent advances in the taxonomy of drosophilids, the high preservation level and the large extension of the sampled fragment and the high complexity of the Atlantic Rain forest. Finally, our data set suggest that the employment of estimators of richness for drosophilid assemblages is useful but it requires caution.

O presente estudo tem como objetivo conhecer a composição da fauna de drosofilídeos em áreas de floresta no sul do Brasil. Além disso, a estimativa da riqueza de espécies desta fauna é brevemente discutida. As amostras foram realizadas em três áreas bem preservadas da Mata Atlântica no estado de Santa Catarina. Neste estudo, 136.931 espécimes foram capturados e 96,6 por cento destes foram identificados em nível específico. A riqueza de espécies observada (153 espécies) é a maior já registrada em inventários faunísticos realizados no Brasil. Dentre as espécies capturadas, sessenta e três não se adequaram às descrições disponíveis e a maioria destas provavelmente não foi descrita. Os estimadores baseados em incidência de espécies tenderam a gerar as mais altas estimativas de riqueza enquanto aqueles baseados em abundância geraram as menores. Tais estimadores sugerem a presença de 172,28 a 220,65 espécies na área estudada. Baseando-se nestes valores, de 69,35 a 88,81 por cento da riqueza de espécies esperada foi amostrada. Sugere-se que a alta riqueza registrada neste estudo é uma conseqüência do grande esforço amostral, do método de captura, de recentes avanços na taxonomia de drosofilídeos, do alto grau de preservação e ampla extensão do fragmento amostrado e da alta complexidade da Mata Atlântica. Finalmente, os dados obtidos sugerem que a aplicabilidade de estimadores de riqueza para assembléias de drosofilídeos é útil, mas requer cautela.