Molecular circumscription of the blackfly Simulium oyapockense from South America.

Problems related to the identity of Simuliidae species are impediments to effective disease control in Amazonia. Some of these species, such as Simulium oyapockense Floch & Abonnenc, 1946 (Diptera: Simuliidae), are vectors of the organisms that cause onchocerciasis and mansonellosis diseases. This blackfly species has a wide distribution in South America, and it is suspected of being a complex of cryptic species. The aim of this study is to characterize the nominal species S. oyapockense using partial COI gene sequences. Seven populations of S. oyapockense (morphologically identified) were analysed, including one from its type-locality. The other six populations were collected in Brazil and in Argentina. A taxon collected in Amazonas state, Brazil, with adults similar to S. oyapockense but with distinct pupae, was also included in the analysis (Simulium 'S'). The nominal species S. oyapockense is circumscribed, and its geographical distribution is restricted to areas north of the Amazon River. Populations of S. oyapockense s.l. collected south of the Amazon River comprise a species complex that needs to be evaluated using integrative taxonomy. Simulium 'S' represents a species with unique morphological and molecular characteristics. Distinguishing cryptic species is a prerequisite for reducing the taxonomic impediment, especially in medically important taxa.

Back from the past: Molecular and morphological support for Simulium mutucuna Nunes de Mello & Vieira da Silva, 1974 (Diptera: Simuliidae) as a valid species.

Simulium mutucuna, a species described based on a single female from Roraima state, was previously synonymized with Simulium paynei and currently is considered a synonym of Simulium rubrithorax. In the present paper we present morphological and molecular evidence supporting the validity of S. mutucuna based on analysis of specimens from Brazil, Venezuela and Mexico. We redescribe the female and describe, for the first time, the male, pupa and larva of S. mutucuna and discuss the morphological differences between this species and the others that are already considered as its senior synonyms. Currently, the distribution of S. mutucuna is restricted to Roraima state. The distribution record for S. rubrithorax in Brazil's North region needs to be removed, since the previous records were based on occurrence of S. mutucuna. Finally, we present new evidence of cryptic diversity in the S. paynei complex based on molecular information.

Description of the immature stages of the Neotropical whirligig beetle Gyrinus (Neogyrinus) rozei Ochs, 1953 (Coleoptera: Gyrinidae) and first report of the parasitoid wasp Melanosmicra sp. (Hymenoptera: Chalcididae) on a Gyrinus species.

In most species of Gyrinidae, the immature stages are unknown, especially due to the difficulty in collecting the juveniles and assigning them to a particular species. Molecular association is a feasible technique that may solve this problem. Recent studies have used DNA sequence data, specifically the gene cytochrome oxidase subunit I (COI), to associate immature and adult stages, thus enabling the description of the former. The objectives of this study were (1) to describe and illustrate the immature stages of Gyrinus (Neogyrinus) rozei Ochs, 1953 including morphometric, chaetotaxic and bionomic information, and (2) to assess the usefulness of the gene COI to associate immatures and adults of Gyrinus. The studied specimens were collected in Roraima state, northern Brazil. The association of immature and adult stages was done either by rearing adults under laboratory conditions or by using DNA sequence data (COI). Eggs were described based on scanning electron microscopy; they are distinguished mainly by several features of chorion, micropyle and reticulation. Larvae of G. (N.) rozei can be distinguished from those of other Neotropical Gyrinidae by a combination of several characters, including the stipes with five robust hook-like additional setae on the dorsointernal margin, and the lacinia roughly hook-shaped. The pupa is similar to that of G. argentinus Steinhel, 1869, except for the number of setae on the body. The first record of the parasitoid wasp Melanosmicra sp. (Hymenoptera: Chalcididae) on a Gyrinus species is also provided.

A New Anthropophilic Species of Simulium (Trichodagmia) (Diptera: Simuliidae) From Amazonia: Morphology, Chromosomes, and DNA Sequences.

The black fly Simulium (Trichodagmia) hirtipupa Lutz (Diptera: Simuliidae) is widely distributed in southern Brazil, with one report from Amapá state in the northern region of Brazilian Amazonia. Morphological comparison of northern and southern populations revealed differences in all life stages, corroborated by chromosomal and molecular analyses, and indicated that the population previously identified as S. hirtipupa from Amapá state represents an undescribed species. This new species is described based on all life stages above the egg, and its chromosomal and molecular divergence from S. hirtipupa is highlighted. Simulium criniferum n. sp. can be diagnosed by the deeply concave male ventral plate with a prominent median projection bearing a ventral keel; female anal lobe in lateral view with a broadly rounded, distal membranous area about as long as wide; pupa with a boot-shaped cocoon bearing a minutely bubbled surface, cephalic plate and thorax with abundant hair-like tubercles, and gill of 12 translucent filaments with darkly sclerotized, acuminate tips; and larva with the body cuticle bearing spiniform setae, abdomen truncated posteriorly, and gill histoblast in situ with the filament tips directed ventrally. Chromosomally, the new species has five unique fixed inversions and uniquely shares three additional fixed inversions with its nearest relative, S. hirtipupa. Partial COI sequences indicate a genetic distance of ~9% between the new species and S. hirtipupa. Females of the new species are anthropophilic.

Not endemic after all: Imparipecten Freeman, 1961 (Diptera: Chironomidae) described from the Neotropical Region.

Imparipecten, a previously monotypic genus, was considered endemic to Australia. Here, we report Imparipecten from the Neotropical region for the first time and describe Imparipecten sychnacanthus sp. n. from Brazil. The association between larvae and adults was established by sequencing a fragment of one ribosomal gene (28S), two fragments of a nuclear protein-coding gene (CAD1 and CAD4), and one mitochondrial protein-coding gene (COI). We also show the close molecular proximity with Imparipecten pictipes through analyses of genetic distances and Bayesian phylogenetics.

Gyretes Brullé (Coleoptera: Gyrinidae) from Brazil: Morphology of eggs and early instars.

Taxonomic information regarding Gyrinidae is mostly based on adults, especially due to the difficulty in collecting immatures and assigning them to a particular species. Association between immatures and adults is sometimes difficult because closely related species can be found in the same habitat. To solve this problem a feasible technique is rearing under laboratory conditions. However, this method is challenging because larval survival rate is usually low, and emulation of natural conditions is difficult. Molecular techniques, especially the use of the COI gene, have been applied to identify species and to associate different life stages. However, in some species groups this marker has not been successful in distinguishing closely related species. The objectives of this study are to describe the egg and the first two instars of Gyretes nubilus Ochs, 1965 and the egg of G. minax Ochs, 1967 and to evaluate the utility of COI to associate immatures and adults. The association of these immature stages with adults was done either rearing adults under laboratory conditions or by using DNA sequence data (COI), corroborating the utility of this molecular marker to associate immature and adults in Gyretes. These immature stages are described, including chaetotaxic analysis of larvae for the first time for the genus Gyretes Brullé, 1835. The eggs are described based on scanning electron microscopy. The eggs are similar to those of other Gyrinidae genera in having a micropylar region in the anterior pole and a longitudinal fissure, and by the absence of an aeropyle, but they differ mainly in characters related to chorionic structure and reticulation. Larvae of Gyretes can be distinguished from those of the other Neotropical Gyrinidae genera by a combination of several characters, including the frontoclypeal seta FR3 short, presence of three conspicuous additional setae on lateral region of parietal (contiguous to stemmata), and posterior margin of lacinia smooth, with apex not indented.

Phylogeography and population diversity of Simulium hirtipupa Lutz (Diptera: Simuliidae) based on mitochondrial COI sequences.

High morphological homogeneity and cryptic speciation may cause the diversity within Simuliidae to be underestimated. Recent molecular studies on population genetics and phylogeography have contributed to reveal which factors influenced the diversity within this group. This study aimed at examining the genetic diversity of Simulium hirtipupa Lutz, 1910 in populations from the biomes Caatinga, Cerrado, and Atlantic Forest. In this study, we carried out phylogeographic and population genetic analyses using a fragment of the mitochondrial gene COI. The 19 populations studied were clustered into seven groups, most of which are associated with geography indicating certain genetic structure. The northern region of the state of Minas Gerais is most likely the center of origin of this species. The average intergroup genetic distance was 3.7%, indicating the presence of cryptic species. The species tree as well as the haplotype network recovered all groups forming two major groups: the first comprises groups Gr-Bahia (in which the São Francisco river has not acted as geographical barrier), Gr-Pernambuco, and Gr-Mato Grosso do Sul. The second included groups comprising populations of the states of Goiás, Tocantins, Minas Gerais, Bahia, São Paulo, and Espírito Santo. The mismatch distribution for groups was consistent with the model of demographic expansion, except for the Gr-Central-East_1 group. The diversification in this group occurred about 1.19 Mya during the Pleistocene, influenced by paleoclimatic oscillations during the Quaternary glacial cycles.

Cytogenetic data on six leafcutter ants of the genus Acromyrmex Mayr, 1865 (Hymenoptera, Formicidae, Myrmicinae): insights into chromosome evolution and taxonomic implications.

Cytogenetic data for the genus Acromyrmex Mayr, 1865 are available, to date, for a few species from Brazil and Uruguay, which have uniform chromosome numbers (2n = 38). The recent cytogenetic data of Acromyrmex striatus (Roger, 1863), including its banding patterns, showed a distinct karyotype (2n = 22), similar to earlier studied Atta Fabricius, 1804 species. Karyological data are still scarce for the leafcutter ants and many gaps are still present for a proper understanding of this group. Therefore, this study aimed at increasing cytogenetic knowledge of the genus through the characterization of other six species: Acromyrmex balzani (Emery, 1890), Acromyrmex coronatus Fabricius, 1804, Acromyrmex disciger (Mayr, 1887), Acromyrmex echinatior (Forel, 1899), Acromyrmex niger (Smith, 1858) and Acromyrmex rugosus (Smith, 1858), all of which were collected in Minas Gerais - Brazil, except for Acromyrmex echinatior which was collected in Barro Colorado - Panama. The number and morphology of the chromosomes were studied and the following banding techniques were applied: C-banding, fluorochromes CMA3 and DAPI, as well as the detection of 45S rDNA using FISH technique. All the six species had the same chromosome number observed for already studied species, i.e. 2n = 38. Acromyrmex balzani had a different karyotype compared with other species mainly due to the first metacentric pair. The heterochromatin distribution also showed interspecific variation. Nevertheless, all the studied species had a pair of bands in the short arm of the first subtelocentric pair. The fluorochrome CMA3 visualized bands in the short arm of the first subtelocentric pair for all the six species, while Acromyrmex rugosus and Acromyrmex niger also demonstrated in the other chromosomes. The AT-rich regions with differential staining using DAPI were not observed. 45S ribosomal genes were identified by FISH in the short arm of the first subtelocentric pair in Acromyrmex coronatus, Acromyrmex disciger and Acromyrmex niger. The uniform chromosome number in the genus Acromyrmex (2n = 38) suggests that Acromyrmex striatus (2n = 22) should be transferred to a new genus. Other aspects of the chromosome evolution in ants are also discussed.

An Asiatic Chironomid in Brazil: morphology, DNA barcode and bionomics.

In most freshwater ecosystems, aquatic insects are dominant in terms of diversity; however, there is a disproportionately low number of records of alien species when compared to other freshwater organisms. The Chironomidae is one aquatic insect family that includes some examples of alien species around the world. During a study on aquatic insects in Amazonas state (Brazil), we collected specimens of Chironomidae that are similar, at the morphological level, to Chironomuskiiensis Tokunaga and Chironomusstriatipennis Kieffer, both with distributions restricted to Asia. The objectives of this study were to provide morphological information on this Chironomus population, to investigate its identity using DNA barcoding and, to provide bionomic information about this species. Chironomus DNA barcode data were obtained from GenBank and Barcode of Life Data Systems (BOLD) and, together with our data, were analyzed using the neighbor-joining method with 1000 bootstrap replicates and the genetic distances were estimated using the Kimura-2-parameter. At the morphological level, the Brazilian population cannot be distinguished either from Chironomusstriatipennis or Chironomuskiiensis, configuring a species complex but, at the molecular level our studied population is placed in a clade together with Chironomusstriatipennis, from South Korea. Bionomic characteristics of the Brazilian Chironomus population differ from the ones of Chironomuskiiensis from Japan, the only species in this species complex with bionomic information available. The Brazilian Chironomus population has a smaller size, the double of the number of eggs and inhabits oligotrophic water, in artificial container. In the molecular analysis, populations of Chironomusstriatipennis and Chironomuskiiensis are placed in a clade, formed by two groups: Group A (which includes populations from both named species, from different Asiatic regions and our Brazilian population) and Group B (with populations of Chironomuskiiensis from Japan and South Korea). Genetic distance between the Brazilian population and specimens in Group A suggests that it was recently introduced in Brazil, and that its country of origin is probably South Korea.

Numerical and structural chromosome variation in the swarm-founding wasp Metapolybia decorata Gribodo 1896 (Hymenoptera, Vespidae).

The Neotropical Polistinae wasps are diverse in taxonomy, social behavior, and nesting founding characteristics. Although some species in this group have been used as models for studies on wasp's biology, they are poorly known in terms of cytogenetics. Here we reported an intraspecific numerical-structural chromosome variation in the swarm-founding wasp Metapolybia decorata from the Brazilian Atlantic Rainforest using conventional and molecular cytogenetic techniques. The observed structural chromosome change involved a telomeric fusion that resulted in a chromosome number range of 2n = 34-36. The origin and geographic distribution of the variant chromosome forms as well as their frequency and maintenance in the studied populations are discussed. In addition, we reported a novel and geographically restricted deletion in the fused chromosomes indicating that the species is undergoing a continued process of karyotype evolution leading to fused chromosome stabilization by elimination of inactive centromeric sequences. Evidence of differences in the telomeric sequences of this wasp was also found by in situ hybridization using the motif (T2AG2)7 as probe.

Occurrence of pre-nucleolar bodies and 45S rDNA location on the chromosomes of the ant Mycocepurus goeldii (Forel) (Formicidae, Myrmicinae, Attini).

The ant Mycocepurus goeldii (Forel) is known for having a relict karyotype with low chromosome number and the present study help the understanding of this ant cytogenetics by describing the occurrence of pre-nucleolar bodies in their chromosomes using impregnation with silver nitrate (Ag-NOR) and the location of 45S rDNA sites by means of the FISH (fluorescent in situ hybridization) technique. Several spots were observed surrounding all chromosomes when submitted to the Ag-NOR technique. These unusual markings were observed in both chromatids of metaphase and early anaphase chromosomes, and are associated to the presence of pre-nucleolar bodies, allowing the observation of the phenomenon of nucleologenesis. Although recent studies have shown that all chromosomes of M. goeldii exhibit centromeric or pericentromeric markings for the CMA(3) fluorochrome, the FISH technique indicated the presence of 45S rDNA in only one pair of chromosomes that differed in the number of CMA(3) markings observed for this species, pointing that the other markings observed with this fluorochrome do not match the sequences in ribosomal genes.

Karyotype characterization of Trigona fulviventris Guérin, 1835 (Hymenoptera, Meliponini) by C banding and fluorochrome staining: report of a new chromosome number in the genus

Although many species of the genus Trigona have been taxonomically described, cytogenetic studies of these species are still rare. The aim of the present study was to obtain cytogenetic data by conventional staining, C banding and fluorochrome staining for the karyotype characterization of the species Trigona fulviventris. Cytogenetic analysis revealed that this species possesses a diploid chromosome number of 2n = 32, different from most other species of this genus studied so far. This variation was probably due to the centric fusion in a higher numbered ancestral karyotype, this fusion producing the large metacentric chromosome pair and the lower chromosome number observed in Trigona fulviventris. Heterochromatin was detected in the pericentromeric region of the first chromosome pair and in one of the arms of the remaining pairs. Base-specific fluorochrome staining with 4'-6-diamidino-2-phenylindole (DAPI) showed that the heterochromatin was rich in AT base pairs (DAPI+) except for pair 13, which was chromomycin A3 (CMA3) positive indicating an excess of GC base pairs. Our data also suggests that there was variation in heterochromatin base composition.