RESUMO
The Mesembrinellidae (Diptera: Oestroidea) comprise a small group of strictly Neotropical calyptrate flies, with 36 described species. The group has often been treated as a subfamily of Calliphoridae, but there is growing evidence that it corresponds to a distinct Oestroidea lineage. Internal relationships have so far been addressed based only on morphology, with results lacking resolution and support. This is the first molecular phylogeny for the group, which is based on the analyses of 80 terminal taxa (22 mesembrinellid and 28 outgroup species) and 5 molecular markers (ITS2, 28S, COI, COII and 16S). Maximum-parsimony, maximum-likelihood and Bayesian inference methods were used, the latter two with partitioning strategies considering codon position and secondary structure information. Results corroborate the Mesembrinellidae as a monophyletic lineage inside Oestroidea. Three clades were consistently recovered: (1) (Laneella + Mesembrinella patriciae); (2) (Mesembrinella (excluding M. patriciae) + Eumesembrinella); and (3) (Huascaromusca + Giovanella). Re-examination of the female reproductive tract of M. patriciae revealed a Laneela-type spermatheca, which corroborates the position of the species recovered in the molecular phylogenetic analyses. Mesembrinella and Huascaromusca are in all cases paraphyletic with regards to Eumesembrinella and Giovanella, respectively. These latter two genera should, thus, be seen as subjective junior synonyms.
RESUMO
The genera Cochliomyia and Chrysomya contain both obligate and saprophagous flies, which allows the comparison of different feeding habits between closely related species. Among the different strategies for comparing these habits is the use of qPCR to investigate the expression levels of candidate genes involved in feeding behavior. To ensure an accurate measure of the levels of gene expression, it is necessary to normalize the amount of the target gene with the amount of a reference gene having a stable expression across the compared species. Since there is no universal gene that can be used as a reference in functional studies, candidate genes for qPCR data normalization were selected and validated in three Calliphoridae (Diptera) species, Cochliomyia hominivorax Coquerel, Cochliomyia macellaria Fabricius, and Chrysomya albiceps Wiedemann . The expression stability of six genes ( Actin, Gapdh, Rp49, Rps17, α -tubulin, and GstD1) was evaluated among species within the same life stage and between life stages within each species. The expression levels of Actin, Gapdh, and Rp49 were the most stable among the selected genes. These genes can be used as reliable reference genes for functional studies in Calliphoridae using similar experimental settings.
Assuntos
Dípteros/genética , Expressão Gênica , Proteínas de Insetos/genética , Animais , Dípteros/crescimento & desenvolvimento , Dípteros/metabolismo , Evolução Molecular , Feminino , Proteínas de Insetos/metabolismo , Larva/genética , Larva/metabolismo , Masculino , Dados de Sequência Molecular , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
Altered acetylcholinesterase (AChE) has been identified in numerous arthropod species resistant to organophosphate (OP) and carbamate insecticides. The New World screwworm (NWS) Cochliomyia hominivorax (Coquerel), one of the most important myiasis-causing flies in the Neotropics, has been controlled mainly by the application of OP insecticides in its current geographical distribution. However, few studies have investigated insecticide resistance in this species. Based on previous studies about mutations conferring OP resistance in related dipteran species, AChE cDNA was sequenced allowing a survey for mutations (I298V, G401A, F466Y) in NWS populations. In addition, the G137D mutation in the carboxylesterase E3 gene, also associated with OP resistance, was analyzed in the same NWS populations. Only 2/135 individuals presented an altered AChE gene (F466Y). In contrast, a high frequency of the G137D mutation in the E3 gene was found in some localities of Brazil and Uruguay, while the mutant allele was not found in Cuba, Venezuela or Colombia. These findings suggest that the alteration in the carboxylesterase E3 gene may be one of the main resistance mechanisms selected in this ectoparasite. The knowledge of the frequency of these resistance-associated mutations in the NWS natural populations may contribute to the selection of appropriate chemicals for control as part of pest management strategies.
Assuntos
Acetilcolinesterase/genética , DNA Complementar/genética , Dípteros/genética , Resistência a Inseticidas/genética , Inseticidas/farmacologia , Organofosfatos/farmacologia , Sequência de Aminoácidos , Animais , Dados de Sequência Molecular , MutaçãoRESUMO
The New World Screwworm (NWS) fly Cochliomyia hominivorax is one of most important myiasis-causing flies in the Neotropics. It is responsible for severe losses to the livestock industry through both mortality and the loss of productivity of infested animals. In Uruguay, NWS represents a significant problem. To date this pest has been controlled by the application of chemical insecticides, mainly the pyrethroid and organophosphate (OP) classes. However, the intensive use of these compounds over many years has led to the evolution of resistance which has the potential to compromise the effectiveness of current control strategies. One mechanism by which resistance has occurred in this and related dipteran species is through two mutations (G137D and W251S) in the carboxylesterase E3 enzyme that have enhanced ability to hydrolyze certain insecticides. In this study changes in the frequency of these mutations in C. hominivorax was investigated in three different Uruguayan regions in 2003 and 2009. All three regions analyzed showed a reduction in the frequency of the G137D mutation and a significant increase in frequency of the W251S mutation, and this may be related to the current intense use of dimethyl-OP and pyrethroid insecticides. The findings of this study provide current information on the frequency of these resistance-associated mutations in NWS in Uruguay and may help select appropriate chemicals for NWS control as part of potential pest management strategies.
Assuntos
Dípteros/enzimologia , Dípteros/genética , Animais , Genótipo , Controle de Insetos , Resistência a Inseticidas/genética , Inseticidas/farmacologia , Mutação , UruguaiRESUMO
The investigation of the genetic variation and population structure of Chrysomya species is of great interest for both basic and applied research. However, very limited genetic information is available for this genus across its geographical distribution. Here, we describe 12 polymorphic microsatellite loci isolated from Chrysomya putoria with expected heterozygosities ranging from 0.1402-0.8312. These markers are of potential applied interest for forensic entomologists and for the characterisation of the genetic structure of C. putoria from recently colonised regions, with great promise for understanding the colonisation dynamics and spread of the genus Chrysomya in the New World.
Assuntos
Dípteros/genética , Repetições de Microssatélites/genética , Polimorfismo Genético/genética , Animais , Brasil , Dípteros/classificação , Frequência do Gene/genética , Marcadores Genéticos/genética , Genética Populacional , Heterozigoto , Análise de Sequência de DNARESUMO
In this report, we describe the development of 17 polymorphic microsatellite markers for the human botfly, Dermatobia hominis, an obligatory parasite of mammals of great veterinary importance in Latin America. The number of alleles ranged from 5 to 21 per locus, with a mean of 12.2 alleles per locus. The expected heterozygosity ranged from 0.2571 to 0.9206 and from 0.2984 to 0.9291 in two populations from Brazil. These markers should provide a high resolution tool for assessment of the fine-scale genetic structure of natural populations of the human botfly.
RESUMO
Chrysomya albiceps is a blowfly of great medical, sanitary and forensic importance widely distributed in the Afrotropical, southern Palaearctic, northern Oriental regions and, recently, in Central and South Americas. Here, we report the characterization of 13 polymorphic microsatellite markers for C. albiceps. The number of alleles ranged from three to 13 alleles with expected heterozygosities ranging from 0.4668 to 0.8408. These markers will be extremely useful for investigating many important aspects of this species such as population structure, dispersal and colonization dynamics.
RESUMO
The horn fly, Haematobia irritans (L.) (Diptera: Muscidae), is a cosmopolitan livestock pest that has caused a great negative impact on the animal production sector throughout the world. Here, we describe 10 polymorphic microsatellite loci isolated from H. irritans. The number of alleles found ranged from two to eight per locus and the expected heterozygosity from 0.1421 to 0.7702. These loci are potentially useful for the fine-scale genetic characterization of horn fly populations and provide fundamental information for pest management and planning of control programs.
RESUMO
Cochliomyia hominivorax (Calliphoridae) is one of the most important myiasis-causing flies and is responsible for severe economic losses to the livestock industry throughout the Neotropical region. In Brazil, C. hominivorax has been controlled mainly with organophosphate (OP) insecticides, although the inappropriate use of these chemicals can result in the selection of resistant flies. Changes in carboxylesterase activity have been associated with OP insecticides in some arthopodan species. In this work, we isolated and characterized part of the E3 gene in C. hominivorax (ChalphaE7), which contained the same substitutions responsible for the acquisition of OP hydrolase activity in Lucilia cuprina (Calliphoridae). Digestion of the polymerase chain reaction products with a restriction enzyme that specifically recognized the mutation site unambiguously differentiated wild and mutated esterase alleles. The PCR-RFLP assay therefore provided a fast, reliable DNA-based method for identifying C. hominivorax individuals with a mutation in the esterase gene. Further bioassays to determine the association of this mutation with OP resistance in C. hominivorax should allow the development of more effective strategies for managing this species.
Assuntos
Dípteros/enzimologia , Resistência a Medicamentos/genética , Esterases/genética , Reação em Cadeia da Polimerase/veterinária , Polimorfismo de Fragmento de Restrição , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Brasil , Esterases/química , Amplificação de Genes , Mutação , Reação em Cadeia da Polimerase/métodos , Mapeamento por Restrição/veterinária , Alinhamento de SequênciaRESUMO
"Myiasis-causing flies" is a generic term that includes species from numerous dipteran families, mainly Calliphoridae and Oestridae, of which blowflies, screwworm flies and botflies are among the most important. This group of flies is characterized by the ability of their larvae to develop in animal flesh. When the host is a live vertebrate, such parasitism by dipterous larvae is known as primary myiasis. Myiasis-causing flies can be classified as saprophagous (free-living species), facultative or obligate parasites. Many of these flies are of great medical and veterinary importance in Brazil because of their role as key livestock insect-pests and vectors of pathogens, in addition to being considered important legal evidence in forensic entomology. The characterization of myiasis-causing flies using molecular markers to study mtDNA (by RFLP) and nuclear DNA (by RAPD and microsatellite) has been used to identify the evolutionary mechanisms responsible for specific patterns of genetic variability. These approaches have been successfully used to analyze the population structures of the New World screwworm fly Cochliomyia hominivorax and the botfly Dermatobia hominis. In this review, various aspects of the organization, evolution and potential applications of the mitochondrial genome of myiasis-causing flies in Brazil, and the analysis of nuclear markers in genetic studies of populations, are discussed.