RESUMO
Bees (Apidae), of which there are more than 19 900 species, are extremely important for ecosystem services and economic purposes, so taxon identity is a major concern. The goal of this study was to optimize the DNA barcode technique based on the Cytochrome c oxidase (COI) mitochondrial gene region. This approach has previously been shown to be useful in resolving taxonomic inconsistencies and for species identification when morphological data are poor. Specifically, we designed and tested new primers and standardized PCR conditions to amplify the barcode region for bees, focusing on the corbiculate Apids. In addition, primers were designed to amplify small COI amplicons and tested with pinned specimens. Short barcode sequences were easily obtained for some Bombus century-old museum specimens and shown to be useful as mini-barcodes. The new primers and PCR conditions established in this study proved to be successful for the amplification of the barcode region for all species tested, regardless of the conditions of tissue preservation. We saw no evidence of Wolbachia or numts amplification by these primers, and so we suggest that these new primers are of broad value for corbiculate bee identification through DNA barcode.
Assuntos
Abelhas/classificação , Abelhas/genética , Código de Barras de DNA Taxonômico/métodos , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Animais , Primers do DNA/genética , DNA Mitocondrial/química , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Análise de Sequência de DNARESUMO
Though the replacement of European bees by Africanized honey bees in tropical America has attracted considerable attention, little is known about the temporal changes in morphological and genetic characteristics in these bee populations. We examined the changes in the morphometric and genetic profiles of an Africanized honey bee population collected near where the original African swarms escaped, after 34 years of Africanization. Workers from colonies sampled in 1968 and in 2002 were morphometrically analyzed using relative warps analysis and an Automatic Bee Identification System (ABIS). All the colonies had their mitochondrial DNA identified. The subspecies that mixed to form the Africanized honey bees were used as a comparison for the morphometric analysis. The two morphometric approaches showed great similarity of Africanized bees with the African subspecies, Apis mellifera scutellata, corroborating with other markers. We also found the population of 1968 to have the pattern of wing venation to be more similar to A. m. scutellata than the current population. The mitochondrial DNA of European origin, which was very common in the 1968 population, was not found in the current population, indicating selective pressure replacing the European with the African genome in this tropical region. Both morphometric methodologies were very effective in discriminating the A. mellifera groups; the non-linear analysis of ABIS was the most successful in identifying the bees, with more than 94% correct classifications.
Assuntos
Abelhas/genética , Animais , Abelhas/anatomia & histologia , Abelhas/classificação , DNA Mitocondrial/genética , Genética Populacional , TempoRESUMO
Mitochondrial genotypes of Africanized honeybees from Brazil and Uruguay were surveyed by DraI restriction of the COI-COII region. Eleven mitotypes were found, three of which had not previously been described (A28-A30). Out of 775 samples (725 from Brazil, 50 from Uruguay), 197 were A1 and 520 were A4. A1 frequency increases toward the north of Brazil, whereas A4 frequency increases toward the south, a pattern echoing the African distribution. The origin of the A4 and most of the A1 African patterns can be attributed to the introduction of Apis mellifera scutellata into Brazil in 1956. The A29 and A30 patterns have the P1 sequence observed in many Iberian Peninsula samples, which represent the traces of the introductions into Brazil and Uruguay by settlers.
Assuntos
Abelhas/enzimologia , Abelhas/genética , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , África , Animais , Sequência de Bases , Brasil , Fluxo Gênico , Genética Populacional , Dados de Sequência Molecular , Polimorfismo de Fragmento de Restrição , Homologia de Sequência do Ácido Nucleico , UruguaiRESUMO
Cytoplasmic line 2 (CL2) has been previously reported as a cytoplasmically inherited chlorophyll-deficient mutant selected from a chloroplast-mutator genotype of barley. It was characterized by a localized effect on the upper part of the first-leaf blade. At emergence the CL2 seedlings-phenotype varied from a grainy light green to an albino color. They gradually greened during the following days, starting from the base of the blade and extending to cover most of its surface when it was fully grown. The present results, from both light microscopy and transmission electron microscopy (TEM), confirmed the previously described positional and time-dependent expression of the CL2 syndrome along the first-leaf blade. During the first days after emergence, light microscopy showed a normally developed chloroplast at the middle part of the CL2 first-leaf blade, meanwhile at the tip only small plastids were observed. TEM showed that the shapes and the internal structure of the small plastids were abnormal, presenting features of proplastids, amyloplasts and/or senescent gerontoplasts. Besides, they lack plastid ribosomes, contrasting with what was observed inside chloroplasts from normal tips, which presented abundant ribosomes. Phenotypic observations and spectrophotometric analysis of seedlings produced by mother plants that had been grown under different temperatures indicated that higher temperatures during seed formation were negatively associated with pigment content in CL2 seedlings. In contrast, higher temperatures during the growth of CL2 seedlings have been associated with increased pigment content. Aqueous solution with kanamycin and streptomycin, which are antibiotics known to interfere with plastid gene translation, were used for imbibition of wild-type and CL2 seeds. Antibiotic treatments differentially reduced the chlorophyll content in the upper part of the first-leaf blade in CL2, but not in wild-type seedlings. These results suggest that in the wild-type, plastid-gene proteins which are necessary for chloroplast development and chlorophyll synthesis in the upper part of the first-leaf blade are usually synthesized during embryogenesis. However, under certain circumstances, in CL2 seedlings, they would be synthesized after germination. In addition, a shortening of the sheath has been observed in association with pigment decrease suggesting the existence of plastid factors affecting the expression of some nuclear genes. We consider the CL2 mutant a unique experimental material useful to study biological phenomena and external factors regulating plastid, and nuclear gene expression during embryogenesis and early seedling development.
Assuntos
Citoplasma/genética , Hordeum/ultraestrutura , Mutação , Hordeum/genética , Hordeum/crescimento & desenvolvimentoRESUMO
The analysis of phenotypic divergence among local populations within a species has been traditionally performed in a spatial context, although advances in genetic analysis using mtDNA have permitted a simultaneous evaluation of geographical and historical patterns of variation, so-called phylogeographical analysis. In this paper, we combine these two dimensions of variation (geographical space and phylogenetic history) to evaluate patterns of phenotypic evolution in honey bees (Apis mellifera L.). Data on 39 phenotypic traits, derived from 417 colonies grouped into 14 subspecies, were analysed using autocorrelation methods. Mantel tests indicated that the relationship between phenotypic divergence, estimated by Euclidean distances among subspecies' morphological centroids, was significant both when compared to geographical distance (r=0.371; P < 0.01) and to genetic distance (estimated as sequence divergence (%) in a mtDNA region encompassing part of the NADH dehydrogenase subunit 2 and isoleucine transfer RNA (r=0.329; P < 0.01)). For the analysis of each trait, the effects of the geographical co-ordinates (latitude and longitude of subspecies geographical range) and of the phylogenetic patterns (defined by eigenvectors of the genetic distance matrix) on phenotypic variation were simultaneously analysed using an extension of a recently developed model, called Phylogenetic Eigenvector Regression (PVR). In general terms, the partial regression slopes indicated that the variation in the characters traditionally associated with adaptive processes, such as body and wing size, were better explained by geographical position. However, characters usually thought to be neutral, such as wing venation angle, were more associated with phylogeny. This is expected because PVR can be interpreted as a partition model, in which adaptive variation tends to be independent of phylogeny (and, in this case, associated with geography). In addition, the first principal component derived from the expected values of the model for each trait, which can be interpreted as the phenotypic variation predicted by phylogeny, is more structured in a north-south cline than are the original data, supporting an adaptive interpretation. The phylogeographical autocorrelation analyses performed in this study show that different traits are more related to one of the two dimensions of variation (geography and phylogeny), and these patterns can furnish insights into the nature of phenotypic evolution in these organisms.