Phylomitogenomics of two Neotropical species of long-legged crickets Endecous Saussure, 1878 (Orthoptera: Phalangopsidae).

Mitochondrial genomes have provided significant insights into the evolution of several insects. A typical mitogenome contains 37 genes, and variations in gene order can indicate evolutionary relationships between species. In this study, we have assembled the first complete mitogenomes of Endecous chape and E. onthophagus and analyzed the phylogenetic implications for the Gryllidea infraorder. We performed DNA extractions and genome sequencing for both Endecous species. Subsequently, we searched for raw data in the Sequence Read Archive (SRA) in NCBI. Using the SRA data, we assembled the partial mitogenome of Dianemobius nigrofasciatus and annotated the protein-coding genes (PCGs) for nine species. Phylogenomic relationships were reconstructed using Maximum Likelihood (ML) and Bayesian Inference (BI), utilizing the PCGs from 49 Gryllidea species. The mitogenome lengths of E. chape and E. onthophagus are 16,266 bp and 16,023 bp, respectively, while D. nigrofasciatus has a length of 15,359 bp. Our results indicate that species within the infraorder exhibit four types of gene order arrangements that align with their phylogenetic relationships. Both phylogenomic trees displayed strong support, and the ML corroborated with the literature. Gryllidea species have significantly contributed to various fields, and studying their mitogenomes can provide valuable insights into this infraorder evolution.

Study of four Neotropical species of tree crickets Oecanthus Serville, 1831 (Orthoptera, Gryllidae) using cytogenetic and molecular markers.

Karyotypes in the worldwide subfamily Oecanthinae show variations in diploid number, chromosome morphology, and sex-chromosome system. This study described the chromosome set and phylogenetic relationships of four Neotropical species, Oecanthus lineolatus, O. valensis, O. pallidus, and O. pictus. We used classical cytogenetics and Bayesian Inference for phylogenetic reconstruction, using the mitochondrial genes COI, 12S rRNA, and 16S rRNA; and analyzed the phylogenetic patterns of changes in chromosome numbers, using ChromEvol. We observed differences in chromosome number among species and two different sex-chromosome systems. Oecanthus pictus showed 2n = 21, X0♂/22, XX♀; O. lineolatus, 2n = 20, XY♂/XX♀; and O. valensis and O. pallidus, 2n = 18, XY♂/XX♀. The karyotype of Oecanthus was asymmetric, one group with large chromosomes and variation in heterochromatin distribution, and another with small acrocentric chromosomes. The phylogenetic tree recovered two main groups: one with the Palearctic species and another with species from different bioregions, but with low posterior probability. The Neotropical species grouped separately, O. valensis and O. pictus with Nearctic and Ethiopian species, and O. pallidus and O. lineolatus in another, well-supported clade. Together, the phylogenic and chromosome data suggest descending dysploidy events during the evolution of the group.

Evolutionary history and classification of Micropia retroelements in Drosophilidae species.

Transposable elements (TEs) have the main role in shaping the evolution of genomes and host species, contributing to the creation of new genes and promoting rearrangements frequently associated with new regulatory networks. Support for these hypotheses frequently results from studies with model species, and Drosophila provides a great model organism to the study of TEs. Micropia belongs to the Ty3/Gypsy group of long terminal repeats (LTR) retroelements and comprises one of the least studied Drosophila transposable elements. In this study, we assessed the evolutionary history of Micropia within Drosophilidae, while trying to assist in the classification of this TE. At first, we performed searches of Micropia presence in the genome of natural populations from several species. Then, based on searches within online genomic databases, we retrieved Micropia-like sequences from the genomes of distinct Drosophilidae species. We expanded the knowledge of Micropia distribution within Drosophila species. The Micropia retroelements we detected consist of an array of divergent sequences, which we subdivided into 20 subfamilies. Even so, a patchy distribution of Micropia sequences within the Drosophilidae phylogeny could be identified, with incongruences between the species phylogeny and the Micropia phylogeny. Comparing the pairwise synonymous distance (dS) values between Micropia and three host nuclear sequences, we found several cases of unexpectedly high levels of similarity between Micropia sequences in divergent species. All these findings provide a hypothesis to the evolution of Micropia within Drosophilidae, which include several events of vertical and horizontal transposon transmission, associated with ancestral polymorphisms and recurrent Micropia sequences diversification.

Combining morphology and molecular data to improve Drosophila paulistorum (Diptera, Drosophilidae) taxonomic status.

The willistoni species subgroup has been the subject of several studies since the latter half of the past century and is considered a Neotropical model for evolutionary studies, given the many levels of reproductive isolation and different evolutionary stages occurring within them. Here we present for the first time a phylogenetic reconstruction combining morphological characters and molecular data obtained from 8 gene fragments (COI, COII, Cytb, Adh, Ddc, Hb, kl-3 and per). Some relationships were incongruent when comparing morphological and molecular data. Also, morphological data presented some unresolved polytomies, which could reflect the very recent divergence of the subgroup. The total evidence phylogenetic reconstruction presented well-supported relationships and summarized the results of all analyses. The diversification of the willistoni subgroup began about 7.3 Ma with the split of D. insularis while D.paulistorum complex has a much more recent diversification history, which began about 2.1 Ma and apparently has not completed the speciation process, since the average time to sister species separation is one million years, and some entities of the D. paulistorum complex diverge between 0.3 and 1 Ma. Based on the obtained data, we propose the categorization of the former "semispecies" of D. paulistorum as a subspecies and describe the subspecies D. paulistorum amazonian, D. paulistorum andeanbrazilian, D. paulistorum centroamerican, D. paulistorum interior, D. paulistorum orinocan and D. paulistorum transitional.

A new species of the genus Rhinoleucophenga (Diptera: Drosophilidae) and redescription of five species from Neotropical region.

The genus Rhinoleucophenga Hendel comprises 29 nominal species with New World distribution. In the present study five species are redescribed: R. angustifrons Malogolowkin; R. lopesi Malogolowkin; R. matogrossensis Malogolowkin; R. nigrescens Malogolowkin and Rhinoleucophenga personata Malogolowkin. R. capixabensis Culik & Ventura is proposed as a new junior synonymy of R. lopesi. Other species, R. jacareacanga sp. nov., is described from the specimens deposited at CEIOC/Fiocruz. The description of new species and review of some former descriptions of Rhinoleucophenga is indispensable since the distribution records of some species are doubtful.

Can sibling species of the Drosophila willistoni subgroup be recognized through combined microscopy techniques?

ABSTRACT In several arthropod groups, male genitalia is the most important feature for species identification, especially in cryptic species. Cryptic species are very common in the Drosophila genus, and the Neotropical Drosophila willistoni species group is a good example. This group currently includes 24 species divided into three subgroups: alagitans, bocainensis and willistoni. There are six sibling species in the willistoni subgroup – D. willistoni, D. insularis, D. tropicalis, D. equinoxialis, D. pavlovskiana and D. paulistorum, which is a species complex composed of six semispecies – Amazonian, Andean-Brazilian, Centroamerican, Interior, Orinocan and Transitional. The objective of this study was to characterize male genitalia of the willistoni subgroup, including the D. paulistorum species complex, using scanning electron microscopy and light microscopy. We also tried to contribute to the identification of these cryptic species and to add some comments about evolutionary history, based on male genitalia characters. Despite being cryptic species, some differences were found among the siblings, including the Drosophila paulistorum semispecies.

The New World genus Rhinoleucophenga (Diptera: Drosophilidae): new species and notes on occurrence records.

The genus Rhinoleucophenga Hendel comprises 26 nominal species with New World distribution. In the present study, two new species are described from samples in the Pampa and Caatinga biomes in Brazil, R. punctuloides sp. nov. and R. trivisualis sp. nov., respectively. Rhinoleucophenga punctuloides sp. nov. is a sibling species of R. punctulata Duda. Furthermore, two females of R. joaquina Schmitz, Gottschalk & Valente were found for the first time and a description is presented. A taxonomic dichotomous key with pictures is given for the Rhinoleucophenga species recorded in the Caatinga and Pampa biomes. The Neotropical open environments are areas of high diversity for Rhinoleucophenga. The description of new species and review of some older descriptions can change the area of species distribution and improve the faunistic knowledge of other localities in which previous studies have shown unidentified or misidentified Rhinoleucophenga species.

Chromosomal evolution in the Drosophila cardini group (Diptera: Drosophilidae): photomaps and inversion analysis.

Detailed chromosome photomaps are the first step to develop further chromosomal analysis to study the evolution of the genetic architecture in any set of species, considering that chromosomal rearrangements, such as inversions, are common features of genome evolution. In this report, we analyzed inversion polymorphisms in 25 different populations belonging to six neotropical species in the cardini group: Drosophila cardini, D. cardinoides, D. neocardini, D. neomorpha, D. parthenogenetica and D. polymorpha. Furthermore, we present the first reference photomaps for the Neotropical D. cardini and D. parthenogenetica and improved photomaps for D. cardinoides, D. neocardini and D. polymorpha. We found 19 new inversions for these species. An exhaustive pairwise comparison of the polytene chromosomes was conducted for the six species in order to understand evolutionary patterns of their chromosomes.

Embryogenesis of Heliconius erato (Lepidoptera, Nymphalidae): a contribution to the anatomical development of an evo-devo model organism.

This study reports on the embryogenesis of Heliconius erato phyllis between blastoderm formation and the prehatching larval stage. Syncytial blastoderm formation occurred approximately 2 h after egg laying (AEL) and at about 4 h, the cellular blastoderm was formed. The germ band arose from the entire length of the blastoderm, and rapidly became compacted occupying approximately two-thirds of the egg length. At about 7 h AEL, protocephalon and protocorm differentiation occurred. Continued proliferation of the germ band was followed by penetration into the yolk mass, forming a C-shaped embryo at about 10 h. Approximately 12 h AEL, the gnathal, thoracic and abdominal segments became visible. The primordium of the mouthparts and thoracic legs formed as paired evaginations, while the prolegs formed as paired lobes. At about 30 h, the embryo reversed dorsoventrally. Approximately 32 h AEL, the protocephalon and gnathal segments fused, shifting the relative position of the rudimentary appendages in this region. At about 52 h, the embryo was U-shaped in lateral view and at approximately 56 h, the bristles began evagination from the larval cuticle. Larvae hatched at about 72 h. We found that H. erato phyllis followed an embryonic pattern consistent with long-germ embryogenesis. Thus, we believe that H. erato phyllis should be classified as a long-germ lepidopteran. The study of H. erato phyllis embryogenesis provided a structural glimpse into the morphogenetic events that occur in the Heliconius egg period. This study could help future molecular approaches to understanding the evolution of Heliconius development.

Reevaluating the infection status by the Wolbachia endosymbiont in Drosophila Neotropical species from the willistoni subgroup.

Infections by the endosymbiotic bacterium Wolbachia developed a rapid global expansion within Old World Drosophila species, ultimately infecting also Neotropical species. In this sense, screenings are necessary to characterize new variants of Wolbachia or new hosts, and also in order to map the dynamics of already known infections. In this paper, we performed a double screening approach that combined Dot-blot and PCR techniques in order to reevaluate the infection status by Wolbachia in species from the willistoni subgroup of Drosophila. Genomic DNA from isofemale lines descendent from females collected in the Amazonian Rainforest (n=91) were submitted to Dot-blot, and were positive for Wolbachia, producing a gradient of hybridization signals, suggesting different infection levels, which was further confirmed through quantitative PCR. Samples with a strong signal in the Dot-blot easily amplified in the wsp-PCR, unlike most of the samples with a medium to weak signal. It was possible to molecularly characterize three Drosophila equinoxialis isofemale lines that were found to be infected in a low density by a wMel-like Wolbachia strain, which was also verified in a laboratory line of Drosophila paulistorum Amazonian. We also found Drosophila tropicalis to be infected with the wAu strain and a Drosophila paulistorum Andean-Brazilian semispecies laboratory line to be infected with a wAu-like Wolbachia. Moreover, we observed that all Drosophila willistoni samples tested with the VNTR-141 marker harbor the same Wolbachia variant, wWil, either in populations from the South or the North of Brazil. Horizontal transfer events involving species of Old World immigrants and Neotropical species of the willistoni subgroup are discussed.

Sex-specific methylation in Drosophila: an investigation of the Sophophora subgenus.

Epigenetic phenomena have been widely characterized in the genomes of vertebrates and DNA methylation is a key mechanism of epigenetic regulation. The DNA methylation systems of invertebrates and vertebrates show several notable differences. However, the evolutionary implications of those differences only recently began to be revealed. Our study investigated the recurrence of sex-specific methylation, as previously described for the species Drosophila willistoni, in other species of the Sophophora subgenus that present close evolutionary relationship. The MSRE and Southern blot techniques were used to analyze rDNA of some species of the willistoni, melanogaster, saltans and obscura groups of Drosophila and the results suggested that differential DNA methylation between sexes only occurs in Drosophila tropicalis and D. insularis, two sibling species of the willistoni subgroup. However, only using the MSRE technique we could detect sex-specific patterns of DNA methylation in all species of willistoni subgroup. These results indicate that DNA methylation may present important differences, even between closely related species, shedding new light on this Neotropical species complex.

The hobo transposon and hobo-related elements are expressed as developmental genes in Drosophila.

Transposable elements comprise a significant part of genomes and are involved in their evolvability. The hobo element is found as an active class II transposable element in Drosophila melanogaster that is able to induce gonadal dysgenesis. Some hobo-related sequences (hRSs) are thought to be relics of old "hobo" invasions, and are therefore ancient genomic constituents. However, some of these hRSs are still mobile. The present study analyzed the expression pattern of hobo and a particular type of hRSs, hobo(VAHS). Both elements were shown to be expressed as sense and antisense mRNA transcripts. Expression analysis in whole mount embryos revealed a pattern similar to that of some developmental regulatory genes. Here we suggest that cis-regulatory sequences similar to those in developmental genes exist in hobo sequences. Therefore, hobo mobilization may contribute to the development of new regulatory networks during genomic evolution.

Over-activation of the Drosophila melanogaster hsp83 gene by selenium intoxication

Selenium is an important dietary micronutrient and an essential component of selenoproteins and the active sites of some enzymes, although at high concentrations it is toxic. We investigated diphenyl diselenide ((C6H5)2Se2 ) for its effects on the developmental stages of Drosophila melanogaster and found that in the larval and pupae stages the toxic threshold for this compound when added to the banana-agar medium on which the larva were fed was 350 µmol. In adult flies, fed on the same media, there were no observable toxic effects below 500 µmol but there were toxic effects above 600 µmol, indicating that adult flies were more resistant to selenium intoxication. In larvae, a when diphenyl diselenide was present above the toxic threshold there was increased activation of the hsp83 heat shock protein gene. Selenium promotes oxidation of sulfhydryl groups and affects the folding of proteins and this could explain the over-expression of hsp83 because the product of this gene is involved in protein folding and defense responses, including the response to heat shock.

First evidence of methylation in the genome of Drosophila willistoni.

DNA methylation has been studied abundantly in vertebrates and recent evidence confirms that this phenomenon could be disseminated among some invertebrates groups, including Drosophila species. In this paper, we used the Methylation-Sensitive Restriction Endonuclease (MSRE) technique and Southern blot with specific probes, to detect methylation in the Drosophila willistoni species. We found differential cleavage patterns between males and females that cannot be explained by Mendelian inheritance, pointing to a DNA methylation phenomenon different from the Drosophila melanogaster one. The sequencing of some of these bands showed that these fragments were formed by different DNA elements, among which rDNA. We also characterized the D. willitoni dDnmt2 sequence, through a Mega Blast search against the D. willistoni Trace Archive Database using the D. melanogaster dDnmt2 nucleotide sequence as query. The complete analysis of D. willistoni dDnmt2 sequence showed that its promoter region is larger, its dDnmt2 nucleotide sequence is 33% divergent from the D. melanogaster one, Inverted Terminal Repeats (ITRs) are absent and only the B isoform of the enzyme is produced. In contrast, ORF2 is more conserved. Comparing the D. willistoni and D. melanogaster dDnmt2 protein sequences, we found higher conservation in motifs from the large domain, responsible for the catalysis of methyl transfer, and great variability in the region that carries out the recognition of specific DNA sequences (TRD). Globally, our results reveal that methylation of the D. willistoni genome could be involved in a singular process of species-specific dosage compensation and that the DNA methylation in the Drosophila genus can have diverse functions. This could be related to the evolutionary history of each species and also to the acquisition time of the dDnmt2 gene.

The genus Zygothrica Wiedemann 1830 (Diptera, Drosophilidae) in Santa Catarina state, southern Brazil: distribution and ecological notes / O gênero Zygothrica Wiedemann 1830 (Diptera, Drosophilidae) no Estado de Santa Catarina, sul do Brasil: distribuição e notas ecológicas

The present paper brings together survey data from nine collection sites in Santa Catarina state, southern Brazil. Samples were obtained during about four years in forested, coastal and urban areas. Flies were attracted by fermented banana baits and captured in traps. Among the species of the genus Zygothrica Wiedemann 1830 collected, Zygothrica apopoeyi Burla 1956, Z. bilineata (Williston 1896), Z. dispar (Wiedemann 1830), Z./ lanceolata Burla 1956, Z. nigropleura Grimaldi 1987, and Z. poeyi (Sturtevant 1921) were recorded for the first time in Santa Catarina state and, except for Z. bilineata, also in southern Brazil. Apart from these species, we also collected Zygothrica hypandriata Burla 1956, Z. orbitalis (Sturtevant 1916) (as Z. parilis), Z. prodispar Duda 1925 and Z. vittimaculosa Burla 1956. Except for Z. bilineata and Z. hypandriata, these are the new southernmost records for the geographical distribution of these species. All the collected species were represented by few individuals, probably as a consequence of the collection method, and most of them was captured in winters and springs. Five species were restricted to the more preserved studied site, and most individuals were collected in forest sites. Nevertheless, four species were also captured in urban or xeric environments, indicating a higher tolerance and a wider ecological versatility in this genus as compared to what was previously thought, at least for some species.

O presente estudo reúne dados de pesquisa de nove locais de coleta no estado de Santa Catarina, sul do Brasil. Foram obtidas amostras durante aproximadamente quatro anos em áreas florestais, litorâneas e urbanas. As moscas foram atraídas através de isca de banana fermentada e capturadas em armadilhas. Dentre as espécies do gênero Zygothrica Wiedemann 1830 coletadas, Zygothrica apopoeyi Burla 1956, Z. bilineata (Williston 1896), Z. dispar (Wiedemann 1830), Z. lanceolata Burla 1956, Z. nigropleura Grimaldi 1987, e Z./poeyi (Sturtevant 1921) foram registradas pela primeira vez no estado de Santa Catarina e, com exceção de Z. bilineata, também no sul do Brasil. Além destas espécies, também foram coletadas Zygothrica hypandriata Burla 1956, Z. orbitalis (Sturtevant 1916) (como Z. parilis), Z. prodispar Duda 1925 e Z. vittimaculosa Burla 1956. Com exceção de Z. bilineata e Z. hypandriata, estes registros representam os novos limites de distribuição geográfica sul para estas espécies. Todas as espécies coletadas foram representadas por poucos indivíduos, provavelmente como conseqüência do método de coleta, e a maioria delas foi capturada em invernos e primaveras. Cinco/espécies foram restritas ao local estudado mais preservado e a maioria dos indivíduos foi coletada em áreas de floresta. Entretanto, quatro espécies também foram capturadas em ambientes urbanos ou xéricos, indicando uma tolerância mais alta e uma versatilidade ecológica mais ampla do que previamente era pensado para este gênero, pelo menos para algumas de suas espécies.

Transposable elements P and gypsy in natural populations of Drosophila willistoni

The presence and integrity of the P transposon and the gypsy retrotransposon in the genome of 18 samples of natural Drosophila willistoni populations collected from a large area of South America were Southern blot screened using Drosophila melanogaster probes. The aim of this screening was provide further knowledge-base on the geographical distribution of D. willistoni and to carry out an inter-population analysis of the P and gypsy elements present in the genomes of the populations analyzed. The fragment patterns obtained indicate that both the P and gypsy elements are ancient in the D. willistoni genome, but whereas the gypsy retroelement appears to be invariable and stable the P element varies between populations and appears to still have some capacity for mobilization.

Chromosomal polymorphism in Drosophila willistoni populations from Uruguay

Chromosomal polymorphism in natural populations of Drosophila willistoni from Uruguay and southern Brazil was investigated in order to understand the genetic characteristics and evolutionary potential of these almost geographically marginal populations. The level of chromosomal polymorphism in samples from Uruguay was higher than in those from the southernmost Brazilian state of Rio Grande do Sul. The increase in the polymorphism of these populations, in which the species almost reaches its southern limit, contradicts the low level of paracentric inversion polymorphism expected under the central-marginal chromosomal polymorphism cline previously reported. The high frequency of some inversions and the presence of unique inversions in samples from Uruguay indicate the uniqueness of these populations