Comparative Cytogenetic Analysis of Three Eumeninae Species (Hymenoptera, Vespidae).

Eumeninae represents the largest subfamily within Vespidae, with 3,600 species described. Of these, only 18 have been cytogenetically analysed. In the present study, we used both classical and molecular techniques to characterise and compare the karyotypes of 3 Eumeninae species, namely, Ancistrocerus sp., Pachodynerus grandis, and Pachodynerus nasidens. Ancistrocerus sp. presented a haploid chromosome number of n = 12, with the first 2 chromosomes of the karyotype being almost entirely heterochromatic and much larger than the remaining chromosomes. The 2 Pachodynerus species presented the same chromosome number (n = 11 and 2n = 22) but displayed different karyotypic formulae. Additionally, chromosomal polymorphisms were observed in the analysed P. nasidens female. In the 3 species, heterochromatin was located in one of the chromosome arms. Fluorochrome staining revealed a balanced composition of AT and GC bases within the chromatin for each of the 3 species, except for few regions that were visibly GC-rich. All species had a single 18S rDNA site that co-localised with GC-rich regions; however, this localisation varied from species to species and not all GC-rich regions corresponded to ribosomal genes. Based on the cytogenetic data obtained here, we discuss the possible numerical/structural rearrangements that may be involved in the karyotypic evolution of the 3 studied species. In addition to the first description of the molecular cytogenetic characteristics of the Eumeninae subfamily and the genus Pachodynerus, this study also provides a relevant contribution towards the discussion of chromosomal evolution in Eumeninae wasps.

Beekeeping practices and geographic distance, not land use, drive gene flow across tropical bees.

Across the globe, wild bees are threatened by ongoing natural habitat loss, risking the maintenance of plant biodiversity and agricultural production. Despite the ecological and economic importance of wild bees and the fact that several species are now managed for pollination services worldwide, little is known about how land use and beekeeping practices jointly influence gene flow. Using stingless bees as a model system, containing wild and managed species that are presumed to be particularly susceptible to habitat degradation, here we examine the main drivers of tropical bee gene flow. We employ a novel landscape genetic approach to analyse data from 135 populations of 17 stingless bee species distributed across diverse tropical biomes within the Americas. Our work has important methodological implications, as we illustrate how a maximum-likelihood approach can be applied in a meta-analysis framework to account for multiple factors, and weight estimates by sample size. In contrast to previously held beliefs, gene flow was not related to body size or deforestation, and isolation by geographic distance (IBD) was significantly affected by management, with managed species exhibiting a weaker IBD than wild ones. Our study thus reveals the critical importance of beekeeping practices in shaping the patterns of genetic differentiation across bee species. Additionally, our results show that many stingless bee species maintain high gene flow across heterogeneous landscapes. We suggest that future efforts to preserve wild tropical bees should focus on regulating beekeeping practices to maintain natural gene flow and enhancing pollinator-friendly habitats, prioritizing species showing a limited dispersal ability.

Cytogenetic analyses using C-banding and DAPI/CMA3 staining of four populations of the maize weevil Sitophiluszeamais Motschulsky, 1855 (Coleoptera, Curculionidae).

Cytogenetic data avalaible for the maize weevil Sitophiluszeamais Motschulsky, 1855 (Coleoptera: Curculionidae), one of the most destructive pests of stored cereal grains, are controversial. Earlier studies focused on single populations and emphasized chromosome number and sex determination system. In this paper, the karyotypes of four populations of Sitophiluszeamais were characterized by conventional staining, C-banding and sequential staining with the fluorochromes chromomycin-A3/4-6-diamidino-2-phenylindole (CMA3/DAPI). The analyses of metaphases obtained from the cerebral ganglia of last instar larvae and the testes of adults showed that the species had 2n = 22 chromosomes, with 10 autosomal pairs and a sex chromosome pair (XX in females and Xyp in males). Chromosome number, however, ranged from 2n = 22 to 26 due to the presence of 0-4 supernumerary chromosomes in individuals from the populations of Viçosa, Unai and Porto Alegre. With the exception of the Y chromosome, which was dot-like, all other chromosomes of this species were metacentric, including the supernumeraries. The heterochromatin was present in the centromeric regions of all autosomes and in the centromere of the X chromosome. The B chromosomes were partially or totally heterochromatic, and the Y chromosome was euchromatic. The heterochromatic regions were labeled with C-banding and DAPI, which showed that they were rich in AT base pairs.

Possible introgression of B chromosomes between bee species (Genus Partamona ).

The origin of supernumerary (B) chromosomes is still a debated topic, with intra- and interspecific origins being the most plausible options. In the bee Partamona helleri, a sequence-characterized amplified region (SCAR) marker being specific to B chromosomes suggested the possibility of interspecific origin. Here, we search for this marker in 3 close relative species and perform DNA sequence comparison between species. The SCAR sequence does not show homology with other sequences in the databases, but does contain an open reading frame with sequence homology with a reverse transcriptase. Dot-blot hybridization using the SCAR marker as a probe confirmed that it is present in B-carrying, but not B-lacking larvae of P. helleri, and indicated its presence in adult individuals of P. cupira and P. criptica. Additionally, PCR amplification of the SCAR marker was successful on genomic DNA obtained from P. helleri and P. rustica larvae carrying B chromosomes, and on genomic DNA obtained from adult individuals of P. cupira, P. criptica and P. rustica. Finally, a comparison of the DNA sequence of the SCAR markers amplified from these 4 species showed very few nucleotide differences between the species. The complete association between B chromosome and SCAR presence and the scarce divergence observed for this DNA sequence between the 4 species analyzed suggest the possibility that this B chromosome has recently been transferred between species through several episodes of interspecific hybridization.

Honey physicochemical properties of three species of the Brazilian Melipona.

Physicochemical analyses were carried out to evaluate 27 samples of honeys from three species of the Brazilian genus Melipona (M. capixaba, M. rufiventris and M. mondury) from Espírito Santo and Minas Gerais States. The parameters water activity (Aw), percentage of soluble solids (Brix %), pH, acidity (meq/Kg) and moisture (%) were evaluated. The honey characteristics obtained from these samples were very similar to the ones from other Melipona species. However, regarding the honey from Apis (honey bee), only the pH values were similar. The low pH value and the high acidity detected in Melipona honey are potential factors for increasing the honey shelf life because they do not provide favorable conditions for the microbial development. On the other hand, the high level of water activity favors the growth of microorganisms, especially yeast, which demands a more careful handled and storage. The observed differences between Melipona and Apis honey reinforce the need for specific quality settings for stingless bee honey.

Transcript levels of ten caste-related genes in adult diploid males of Melipona quadrifasciata (Hymenoptera, Apidae) - A comparison with haploid males, queens and workers.

In Hymenoptera, homozygosity at the sex locus results in the production of diploid males. In social species, these pose a double burden by having low fitness and drawing resources normally spent for increasing the work force of a colony. Yet, diploid males are of academic interest as they can elucidate effects of ploidy (normal males are haploid, whereas the female castes, the queens and workers, are diploid) on morphology and life history. Herein we investigated expression levels of ten caste-related genes in the stingless bee Melipona quadrifasciata, comparing newly emerged and 5-day-old diploid males with haploid males, queens and workers. In diploid males, transcript levels for dunce and paramyosin were increased during the first five days of adult life, while those for diacylglycerol kinase and the transcriptional co-repressor groucho diminished. Two general trends were apparent, (i) gene expression patterns in diploid males were overall more similar to haploid ones and workers than to queens, and (ii) in queens and workers, more genes were up-regulated after emergence until day five, whereas in diploid and especially so in haploid males more genes were down-regulated. This difference between the sexes may be related to longevity, which is much longer in females than in males.

Transcript levels of ten caste-related genes in adult diploid males of Melipona quadrifasciata (Hymenoptera, Apidae): a comparison with haploid males, queens and workers

In Hymenoptera, homozygosity at the sex locus results in the production of diploid males. In social species, these pose a double burden by having low fitness and drawing resources normally spent for increasing the work force of a colony. Yet, diploid males are of academic interest as they can elucidate effects of ploidy (normal males are haploid, whereas the female castes, the queens and workers, are diploid) on morphology and life history. Herein we investigated expression levels of ten caste-related genes in the stingless bee Melipona quadrifasciata, comparing newly emerged and 5-day-old diploid males with haploid males, queens and workers. In diploid males, transcript levels for dunce and paramyosin were increased during the first five days of adult life, while those for diacylglycerol kinase and the transcriptional co-repressor groucho diminished. Two general trends were apparent, (i) gene expression patterns in diploid males were overall more similar to haploid ones and workers than to queens, and (ii) in queens and workers, more genes were up-regulated after emergence until day five, whereas in diploid and especially so in haploid males more genes were down-regulated. This difference between the sexes may be related to longevity, which is much longer in females than in males.

Variation and genetic structure of Melipona quadrifasciata Lepeletier (Hymenoptera, Apidae) populations based on ISSR pattern.

For a study of diversity and genetic structuring in Melipona quadrifasciata, 61 colonies were collected in eight locations in the state of Minas Gerais, Brazil. By means of PCR analysis, 119 ISSR bands were obtained, 80 (68%) being polymorphic. H(e) and H (B) were 0.20 and 0.16, respectively. Two large groups were obtained by the UPGMA method, one formed by individuals from Januária, Urucuia, Rio Vermelho and Caeté and the other by individuals from São João Del Rei, Barbacena, Ressaquinha and Cristiano Otoni. The Φst and θ(B) values were 0.65 and 0.58, respectively, thereby indicating high population structuring. UPGMA grouping did not reveal genetic structuring of M. quadrifasciata in function of the tergite stripe pattern. The significant correlation between dissimilarity values and geographic distances (r = 0.3998; p < 0.05) implies possible geographic isolation. The genetic differentiation in population grouping was probably the result of an interruption in gene flow, brought about by geographic barriers between mutually close geographical locations. Our results also demonstrate the potential of ISSR markers in the study of Melipona quadrifasciata population structuring, possibly applicable to the studies of other bee species.

Variation and genetic structure of Melipona quadrifasciata Lepeletier (Hymenoptera, Apidae) populations based on ISSR pattern

For a study of diversity and genetic structuring in Melipona quadrifasciata, 61 colonies were collected in eight locations in the state of Minas Gerais, Brazil. By means of PCR analysis, 119 ISSR bands were obtained, 80 (68 percent) being polymorphic. He and H B were 0.20 and 0.16, respectively. Two large groups were obtained by the UPGMA method, one formed by individuals from Januária, Urucuia, Rio Vermelho and Caeté and the other by individuals from São João Del Rei, Barbacena, Ressaquinha and Cristiano Otoni. The Φst and θB values were 0.65 and 0.58, respectively, thereby indicating high population structuring. UPGMA grouping did not reveal genetic structuring of M. quadrifasciata in function of the tergite stripe pattern. The significant correlation between dissimilarity values and geographic distances (r = 0.3998; p < 0.05) implies possible geographic isolation. The genetic differentiation in population grouping was probably the result of an interruption in gene flow, brought about by geographic barriers between mutually close geographical locations. Our results also demonstrate the potential of ISSR markers in the study of Melipona quadrifasciata population structuring, possibly applicable to the studies of other bee species.

Genetic variability of native populations of Wasmannia Forel (Hymenoptera: Formicidae) and their biogeographical implications.

Wasmannia auropunctata (Roger) and Wasmannia rochai Forel are economically important ants in the Southeast and Southwest regions of Bahia State, Brazil. Randomly Amplified Polymorphic DNA (RAPD) markers were used to distinguish and analyze the genetic variability of populations of both species from Ilhéus, Jequié, BA and French Guyana. The genetic distances among W. auropunctata and W.rochai populations (55.8-71.4%) suggest genetic differentiation among them. Wasmannia auropunctata populations from sugarcane and banana plantations in Jequié were the most distant genetically (30.1-46.3%) and may represent populations restricted to isolated fragments. The high genetic distances among W. auropunctata populations from CEPLAC experimental areas, in Ilhéus (26.8-34.6%) and the other populations from Ilhéus (23.3-40.8%), suggest a multicolonial structure of W. auropunctata in southeast Bahia. The genetic proximity among the W. auropunctata populations from cocoa (14.1%) and coconut plantations (18.5%) in Ilhéus with the populations from the French Guyanan forests suggest that there was recent and large expansion of populations derived from a single population, that are today distributed in habitats with similar environmental characteristics. The high polymorphism and the estimated heterozygosity values for the two species suggest that we studied native W. auropunctata and W.rochai populations.

Genetic similarity among Agistemus pallinii Matioli et al (Acari: Stigmaeidae) found in citrus orchards in Viçosa, Minas Gerais State, Brazil.

Stigmaeidae are very important predators of mite and insect pests on several crops in Brazil. It is considered the second most important family of predatory mites in citrus orchards in Brazil. However, their identification, especially that of the members of the genus Agistemus, is rather difficult based only on morphology. Hence, this study describes the use of random amplified polymorphic DNA (RAPD-PCR) markers to determine the genetic similarity of an Agistemus pallinii Matioli et al population found in 2004 in a citrus orchard in Viçosa, Minas Gerais State, Brazil, preying on Panonychus citri (McGregor). Amplifi cations were performed with 12 random primers (OPAA8, OPAA19, OPAB1, OPAB5, OPAB18, OPAC9, OPAC17, OPAC19, OPAD10, OPAE9, OPAE12 and OPAE17), which generated 119 bands, with 53.8% polymorphism. The coefficients of genetic similarity among the individuals ranged from 0.68 to 0.99, indicating a high genetic similarity among them. The 3D projection analysis clustered the majority of individuals confirming their high similarity. Though individuals of A. pallinii are minute (+/- 360 microm long), the PCR-RAPD technique can still be used for their identification, complementing morphological analyses or for comparison of populations collected in different geographic regions. This is the first molecular study carried out with stigmaeid mites.

Genetic variability of native populations of Wasmannia Forel (Hymenoptera: Formicidae) and their biogeographical implications / Variabilidade genética de populações nativas de espécies de Wasmannia Forel (Hymenoptera: Formicidae) e suas implicações biogeográficas

Wasmannia auropunctata (Roger) and Wasmannia rochai Forel are economically important ants in the Southeast and Southwest regions of Bahia State, Brazil. Randomly Amplified Polymorphic DNA (RAPD) markers were used to distinguish and analyze the genetic variability of populations of both species from Ilhéus, Jequié, BA and French Guyana. The genetic distances among W. auropunctata and W.rochai populations (55.8-71.4 percent) suggest genetic differentiation among them. Wasmannia auropunctata populations from sugarcane and banana plantations in Jequié were the most distant genetically (30.1-46.3 percent) and may represent populations restricted to isolated fragments. The high genetic distances among W. auropunctata populations from CEPLAC experimental areas, in Ilhéus (26.8-34.6 percent) and the other populations from Ilhéus (23.3-40.8 percent), suggest a multicolonial structure of W. auropunctata in southeast Bahia. The genetic proximity among the W. auropunctata populations from cocoa (14.1 percent) and coconut plantations (18.5 percent) in Ilhéus with the populations from the French Guyanan forests suggest that there was recent and large expansion of populations derived from a single population, that are today distributed in habitats with similar environmental characteristics. The high polymorphism and the estimated heterozygosity values for the two species suggest that we studied native W. auropunctata and W.rochai populations.

Wasmannia auropunctata (Roger) e Wasmannia rochai Forel são duas formigas economicamente importantes no Sudeste e Sudoeste da Bahia. Marcadores de RAPD (polimorfismo de DNA amplificado ao acaso) foram utilizados para distinguir e analisar a variabilidade genética de populações de ambas as espécies, provenientes de Ilhéus e Jequié, BA, e da Guiana Francesa. As distâncias genéticas entre populações de W. auropunctata e W.rochai (55,8-71,4 por cento) evidenciam a diferenciação genética entre elas. As populações de W. auropunctata presentes em canaviais e bananais de Jequié mostraram-se geneticamente mais distantes (30,1-46,3 por cento), podendo representar populações restritas a fragmentos isolados. As elevadas distâncias genéticas entre as populações de W. auropunctata das áreas experimentais da CEPLAC, em Ilhéus (26,8-34,6 por cento), assim como as demais populações de Ilhéus (23,3-40,8 por cento), sugerem uma estrutura multicolonial de W. auropunctata no Sudeste da Bahia. A proximidade genética entre as populações de W. auropunctata dos cacauais (14,1 por cento) e coqueirais de Ilhéus (18,5 por cento) com as populações de floresta da Guiana Francesa sugere que houve expansão grande e recente de populações oriundas de uma única população a partir de um centro não-determinado, estando hoje distribuídas em habitats com características ambientais semelhantes. O elevado polimorfismo e os valores das heterozigosidades estimadas para as duas espécies sugerem que foram analisadas populações nativas de ambas as espécies.

Genetic similarity among Agistemus pallinii Matioli et al (Acari: Stigmaeidae) found in citrus orchards in Viçosa, Minas Gerais State, Brazil / Similaridade genética entre Agistemus pallinii Matioli et al (Acari: Stigmaeidae) encontrados em pomares de citros em Viçosa, MG

Stigmaeidae are very important predators of mite and insect pests on several crops in Brazil. It is considered the second most important family of predatory mites in citrus orchards in Brazil. However, their identification, especially that of the members of the genus Agistemus, is rather difficult based only on morphology. Hence, this study describes the use of random amplified polymorphic DNA (RAPD-PCR) markers to determine the genetic similarity of an Agistemus pallinii Matioli et al population found in 2004 in a citrus orchard in Viçosa, Minas Gerais State, Brazil, preying on Panonychus citri (McGregor). Amplifi cations were performed with 12 random primers (OPAA8, OPAA19, OPAB1, OPAB5, OPAB18, OPAC9, OPAC17, OPAC19, OPAD10, OPAE9, OPAE12 and OPAE17), which generated 119 bands, with 53.8 percent polymorphism. The coefficients of genetic similarity among the individuals ranged from 0.68 to 0.99, indicating a high genetic similarity among them. The 3D projection analysis clustered the majority of individuals confirming their high similarity. Though individuals of A. pallinii are minute (± 360 µm long), the PCR-RAPD technique can still be used for their identification, complementing morphological analyses or for comparison of populations collected in different geographic regions. This is the first molecular study carried out with stigmaeid mites.

Stigmaeidae são importantes predadores de ácaros e insetos pragas em várias culturas no Brasil. Essa família é considerada a segunda mais importante de ácaros predadores em citros no Brasil. Entretanto, o status taxonômico desses ácaros, principalmente do gênero Agistemus, é difícil de ser analisado com base em caracteres morfológicos. Assim, este trabalho descreve o uso de marcadores polimórfi cos amplificados ao acaso (PCR-RAPD) para investigar a similaridade genética de uma população de Agistemus pallinii Matioli et al encontrada em 2004 em pomar de citros em Viçosa, MG, predando Panonychus citri (McGregor). As amplificações foram realizadas com 12 iniciadores randômicos (OPAA8, OPAA19, OPAB1, OPAB5, OPAB18, OPAC9, OPAC17, OPAC19, OPAD10, OPAE9, OPAE12 e OPAE17), os quais geraram 119 bandas, sendo 53,8 por cento polimórficas. Os coeficientes de similaridade genética entre os indivíduos variaram de 0,68 a 0,99, indicando alta similaridade entre eles. A análise de projeção gráfica 3D mostrou que a maioria dos indivíduos analisados formou um único grupo, confirmando a alta similaridade genética entre os mesmos. Assim, mesmo considerando o pequeno tamanho de A. pallinii (± 360 µm), a técnica de PCR-RAPD pode ser utilizada para a sua identificação, em suporte às análises morfológicas ou em comparações de populações coletadas em diferentes regiões geográficas. O presente trabalho representa o primeiro estudo molecular realizado com ácaros da família Stigmaeidae.

RAPD variation within and among species of Melittobia Westwood (Hymenoptera: Eulophidae)

O gênero Melittobia Westwood compreende várias espécies de microparasitóides, das quais apenas duas foram registradas no Brasil até agora: M. australica Girault e M. hawaiiensis Perkins. Entretanto, essas espécies são de difícil identificação através de métodos taxonômicos tradicionais. No presente trabalho, marcadores moleculares amplificados ao acaso (PCR-RAPD) foram utilizados com o objetivo de se discriminar as duas espécies e, ao mesmo tempo, analisar a variabilidade genética em populações de M. australica. A maioria dos fragmentos gerados foi espécie-específicos estando presente em todos os indivíduos de uma espécie e ausente nos indivíduos da outra espécie, demonstrando a adequação dessa técnica na distinção das espécies de Melittobia estudadas. A técnica de PCR-RAPD também demonstrou que os indivíduos das diferentes populações estudadas são muito semelhantes entre si, o que pode ser atribuído ao efeito fundador e/ou a grande capacidade de dispersão dessas populações. As distâncias genéticas dentro (D = 1,19-3,54%) e entre as populações (D = 1,93-5,28%) de M. australica apresentaram valores muito baixos, refletindo a reduzida variação genética existente nas populações dessa espécie.

RAPD variation within and among species of Melittobia westwood (Hymenoptera: Eulophidae).

The genus Melittobia Westwood comprises several species of microparasitoids and only two of them are know to occur in Brazil up to now: M. australica Girault and M. hawaiiensis Perkins. Nevertheless, the differentiation between these two species using traditional taxonomy is very difficult. In the present study, we used random amplified polymorphic DNA chain reaction (RAPD-PCR) to test for its ability to discriminate between these two species and to examine the genetic variation among the studied populations of M. australica. Most of the generated fragments were species-specific, occurring in all individuals of one species and absent in the individuals of the other species demonstrating the appropriateness of such technique to distinguish between both of the Melittobia species occurring in Brazil. RAPD-PCR also demonstrated low variability among different populations of M. australica, which may be due to a founder effect and/or high dispersion capacity of these populations. Genetic distances within (D = 1.19-3.54%) and among populations (D = 1.93-5.28%) presented very low values, reflecting the reduced genetic variation that exists among populations of M. australica.