Ana Maria Lima de Azeredo-Espin

The mitochondrial genome of the primary screwworm fly Cochliomyia hominivorax (Diptera: Calliphoridae)

The complete sequence of the mitochondrial genome of the screwworm Cochliomyia hominivorax was determined. This genome is 16 022 bp in size and corresponds to a typical Brachycera mtDNA. A Serine start codon for COI and incomplete termination codons for COII, NADH 5 and NADH 4 genes were described. The nucleotide composition of C. hominivorax mtDNA is 77% AT‐rich, reflected in the predominance of AT‐rich codons in protein‐coding genes. Non‐optimal codon usage was commonly observed in C. hominivorax mitochondrial genes. Phylogenetic analysis distributed the Acalypterate species as a monophyletic group and assembled the C. hominivorax (Calyptratae) and the Acalyptratae in a typical Brachycera cluster. The identification of diagnostic restriction sites on the sequenced mitochondrial genome and the correlation with previous RFLP analysis are discussed.

Levantamento de microhimenópteros parasitóides de dípteros de importância, médico-veterinária no Brasil

De larvas e pupas de Musca domestica, Chrysomya albiceps, Cochliomyia homivorax, Stomoxys calcitrans e Syntesiomyia nudiseta coletadas em diversos ambiente, em Sao Paulo, Parana, Mato Grosso, Mato Grosso do Sul e Minas Gerais, foram obtidas dez especies de microhimenopteros parasitoides da supermamilia Chalcidoidea, algumas assinaladas pela primeira vez no Brasil.

Evolution and structural organisation of mitochondrial DNA control region of myiasis-causing flies.

This study reports the molecular characterization of the mtDNA control region (called the A + T‐rich region in insects) of five dipteran species which cause myiasis: Cochliomyia hominivorax Coquerel, Cochliomyia macellaria Fabricius, Chrysomya megacephala Fabricius, Lucilia eximia Wiedemann (Diptera: Calliphoridae) and Dermatobia hominis Linnaeus Jr (Diptera: Oestridae). The control region in these species varies in length from 1000 to 1600 bp. Two structural domains with specific evolutionary patterns were identified. These were (1) conserved sequence blocks containing primary sequence motifs, including dinucleotide pyrimidine‐purine series and long T‐stretches, located at the 5′ end adjacent to the tRNAIle gene and (2) a hypervariable domain at the 3′ end characterized by increased nucleotide divergence and size variation. A high frequency of A↔T transversions at nucleotide substitution level indicated directional mutation pressure. The phylogenetic usefulness of the insect control region is discussed.

AMiGA: the arthropodan mitochondrial genomes accessible database

UNLABELLED The Arthropodan Mitochondrial Genomes Accessible database (AMiGA) is a relational database developed to help in managing access to the increasing amount of data arising from developments in arthropodan mitochondrial genomics (136 mitochondrial genomes as of September 2005). The strengths of AMiGA include (1) a more accessible and up-to-date database containing a more comprehensive set of mitochondrial genomes for this phylum, (2) the provision of flexible search options for retrieving detailed information such as bibliographical data, genomic graphics, FASTA sequences and taxonomical status, (3) the possibility of enhanced comparative analyses by multiple alignment of single or concatenated sets of genes, (4) more accurate and updated information resulting from a specific curation process called AMiGA Notes and (5) the possibility of including unpublished sequences in a password-restricted area for comparative analysis with the other sequences stored in the database. AVAILABILITY http://amiga.cbmeg.unicamp.br CONTACT lessinger@amiga.cbmeg.unicamp.br SUPPLEMENTARY INFORMATION Detailed information, including an illustrated tutorial, is available from the above URL.

Structure and evolution of the mitochondrial genomes of Haematobia irritans and Stomoxys calcitrans : The Muscidae (Diptera: Calyptratae) perspective

We present the first two mitochondrial genomes of Muscidae dipterans for the species Haematobia irritans (the horn fly) and Stomoxys calcitrans (the stable fly). Typical insect mtDNA features are described, such as a high A+T content (79.1% and 78.9%, respectively), the preference for A+T-rich codons, and the evidence of a non-optimal codon usage. The strong A+T enrichment partially masks another nucleotide content bias maintained by A+C mutation pressure in these Muscidae mtDNAs. The analysis of this data provides a model of metazoans tRNA anticodon evolution, based on the selection hypothesis of anticodon versatility. H. irritans mitochondrial genome (16078 bp) is structurally similar to the hypothetical ancestral mitochondrial genome of arthropods and its control region (A+ T-rich region in insects) organization is consistent with the structure described for Brachycera dipterans. On the other hand, the mitochondrial genome of S. calcitrans is approximately 2kb longer (18 kb), characterized by the presence of approximately 550 bp tandem repeats in the control region, and an extra copy of trnI remarkably similar to a duplicated element of blowflies mtDNA. Putative sequence elements, involved in the regulation of transcription and replication of the mtDNA, were reliably identified in S. calcitrans control region despite the 0.8-1.5 kb gap uncovered from this genome. The use of amino acid and nucleotide sequences of concatenated mitochondrial protein-coding genes (PCGs) in phylogenetic reconstructions of Diptera does not support the monophyly of Muscomorpha, as well as the monophyly of Acalyptratae. Within the Calyptratae group, the inclusion of Muscidae (Muscoidea) as a sister group of Calliphoridae (Oestroidea) implies in a potential conflict concerning the monophyly of the superfamily Oestroidea.

Molecular phylogenetics of Oestroidea (Diptera: Calyptratae) with emphasis on Calliphoridae: Insights into the inter-familial relationships and additional evidence for paraphyly among blowflies

The superfamily Oestroidea, comprising ∼15,000 species, is a large and ecologically diverse clade within the order Diptera. Among its six commonly recognized families, Calliphoridae seems to be crucial for understanding evolutionary relationships in the group, as it is recognized as a controversial paraphyletic grouping. To further investigate this matter, the ITS2, 28S, COI and 16S regions were used to infer phylogenetic relationships in Oestroidea with maximum-parsimony (MP), maximum-likelihood (ML) and Bayesian inference (BI) methods. For the BI analyses, a deep evaluation of different data partitioning strategies was conducted, including consideration of structural conformation (ITS2 and 16S) and codon position (COI) information. Results suggest the existence of two main clades in Oestroidea: (Tachinidae+Mesembrinellinae) and (Rhiniinae, (Sarcophagidae+Calliphoridae sensu stricto)). Oestridae was recovered as sister group of the remaining Oestroidea in the MP trees while it was placed closer to the (Rhiniinae+Sarcophagidae+Calliphoridae sensu stricto) group in the ML and BI trees. A paraphyletic Calliphoridae was recovered, confirming the exclusion of Rhiniinae, a clade recently promoted to family status and therefore already excluded. Mesembrinellinae could also be considered a distinct group apart from Calliphoridae, although further studies are required. Consideration of structural and codon position information led to a significant increase in the log-likelihoods of the analyses, which were accompanied by small changes in the inferred topologies, branch lengths and posterior probability support values. However, as model complexity increases, so does uncertainty across the estimated parameters, including tree topologies, and phylogenies inferred under very parameter-rich models may be less reliable even when possessing higher log-likelihoods.

Genetic Approaches for Studying Myiasis-causing Flies: Molecular Markers and Mitochondrial Genomics

Abstract“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.

Detection and Genetic Diversity of a Heliothine Invader (Lepidoptera: Noctuidae) From North and Northeast of Brazil

ABSTRACT The cotton bollworm, Helicoverpa armigera (Hübner), was recently introduced in Brazil. During the 2012-2013 harvest, producers reported reduced yields up to 35% on major crops. The economic losses reached US

Methods for the recovery of mitochondrial DNA sequences from museum specimens of myiasis‐causing flies

1 billion only in western Bahia, triggering a phytosanitary crisis. The deficiencies in existing taxonomic keys to deal with the morphologically indistinct larvae of H. armigera and the native Helicoverpa zea (Boddie) constrained the detection of new incursions of this heliothine invader. This study explored the identity of heliothine larvae that were found infesting soybean- and corn-growing areas from Roraima state, northern Brazil, through sequences of the mitochondrial cytochrome c oxidase subunit I gene. The inter- and intraspecies sequence variations of DNA barcodes in H. armigera and H. zea were analyzed. The genetic diversity and population structure of the specimens from Roraima and two populations from Piaui and Bahia states, northeastern Brazil, were assessed by adding the cytochrome coxidase subunit II gene to the analysis. Owing to the lack of studies on genetic introgression for the two species, the suitability of using three different nuclear genes to distinguish the two species was also investigated. The results showed strong evidence that the heliothine larvae from north and northeast of Brazil are conspecific with H. armigera, suggesting that this invasive moth has already crossed the Amazon basin. Surveys in the north of South America should start as soon as possible to monitor the entry or spread of this moth in the Caribbean, Central America, and the United States.

The Mitochondrial DNA Control Region of Muscidae Flies: Evolution and Structural Conservation in a Dipteran Context

Abstract Mitochondrial DNA (mtDNA) sequences from eight species of myiasis‐causing flies, stored for up to 50 years, were amplified successfully. Universal primers were used to amplify six specific regions from total genomic DNA, including five mtDNA genes. The comparison of phenol/chloroform, DNAzol® and Chelex techniques for DNA extraction showed that the DNAzol® reagent was the most efficient in retrieving DNA from museum specimens, although the Chelex extraction procedure is currently the most frequently reported method. Comparison of the universal primer sequences with the homologous sequences of Cochliomyia hominivorax Coquerel and Chrysomya putoria Wiedemann (Diptera: Calliphoridae) revealed mismatches that could contribute to the low recovery of a short sequence from subunit II of cytochrome oxidase. The ability to characterize mtDNA markers from museum specimens should be useful in comparative studies of contemporary samples and should help in elucidating species introduction, colonization and dispersal.