What's in a child's ear? A case of otomyiasis by Sarcophaga argyrostoma (Diptera, Sarcophagidae).

A clinical report of otomyiasis in a 1-year-old girl is reported. A III instar larva of Sarcophaga sp. was microscopically identified and Sarcophaga (Liopygia) argyrostoma (Diptera, Sarcophagidae) was suspected. A molecular method targeting a fragment of the cox1 gene was used to confirm the identity of the specimen. Although myiases are not frequent manifestations in otolaryngology, they should arouse the attention of doctors, social workers and parents dealing with disabled people, the elderly and children. This contribution also highlights the need of combining microscopy and molecular tools to achieve a correct and reliable identification of the specimen/s.

A phylotranscriptomic framework for flesh fly evolution (Diptera, Calyptratae, Sarcophagidae).

The Sarcophagidae (flesh flies) comprise a large and widely distributed radiation within the Calyptratae (Diptera). Larval feeding habits are ecologically diverse and include sarcosaprophagy, coprophagy, herbivory, invertebrate and vertebrate predation, and kleptoparasitism. To elucidate the geographic origin and evolution of flesh fly life-history, we inferred a backbone phylogeny based on transcriptomic data from 26 sarcophagid species covering all three subfamilies plus 15 outgroups. The phylogeny was inferred using maximum parsimony and maximum likelihood methods based on a series of supermatrices, one set with overall information content improved by MARE (2290 loci), one set with 100% gene coverage for all included species (587 loci), and the last set including mitochondrial and nuclear genes (589 loci) and additional taxa. In order to obtain a more detailed hypothesis, we utilized the supertree approach to combine results from the present study with previously published hypotheses. This resulted supertree covers 84 of the one hundred currently recognized sarcophagid genera and formed the basis for the ancestral state reconstructions. The monophyletic Sarcophagidae is well-supported as sister to {Mystacinobiidae + Oestridae}, and relationships at the subfamily level are inferred as {Sarcophaginae, (Paramacronychiinae + Miltogramminae)}. The Sarcophagidae and each subfamily originated in the Americas, with Sarcophaginae diversifying mainly in the Neotropics, whereas the major radiation of both Miltogramminae and Paramacronychiinae occurred in the Palaearctic. Sarcosaprophagy is reconstructed as the ancestral larval feeding habit of the family Sarcophagidae and each subfamily. The ancestral sarcophagid larva probably utilized dead invertebrates as food, and the food spectrum expanded together with the diversification of breeding strategies. Particularly, kleptoparasitism in Miltogramminae is derived from sarcosaprophagy and may be seen as having derived from the breeding biology of 'lower' miltogrammines, the larvae of which feed on buried vertebrate carrion.

Species Delimitation of Scavenger Flies in the Valley of Mexico.

Identification of species involved in cadaveric decomposition, such as scavenger Diptera, is a fundamental step for the use of entomological evidence in court. Identification based on morphology is widely used in forensic cases; however, taxonomic knowledge of scavenger fauna is poor for many groups and for many countries, particularly Neotropical ones. A number of studies have documented the utility of a DNA barcoding strategy to assist in the identification of poorly known and diverse groups, particularly in cases involving immature states or fragmented organisms. To provide baseline knowledge of the diversity of scavenger Diptera in the Valley of Mexico, we generated a DNA barcode collection comprised of sequences of the cytochrome c oxidase subunit 1 (COI) gene for all families sampled at a nature reserve located in this region. We collected and identified specimens on the basis of morphology and a species delimitation analysis. Our analyses of 339 individuals delineated 42 species distributed across nine families of Diptera. The richest families were Calliphoridae (9 species), Sarcophagidae (7 species), and Phoridae (6 species). We found many of the species previously recorded for the Valley of Mexico, plus 18 new records for the region. Our study highlights the utility of DNA barcoding as a first-step strategy to assess species richness of poorly studied scavenger fly taxa.

Diversity of Necrophagous Flies (Diptera: Calliphoridae, Mesembrinellidae, and Sarcophagidae) in Anthropogenic and Preserved Environments of Five Different Phytophysiognomies in Northeastern Brazil.

The present study investigated the potential association between the richness and abundance of species of the dipteran families Calliphoridae, Mesembrinellidae, and Sarcophagidae in anthropogenic and preserved environments of five phytophysiognomies (Cerrado, Amazon forest, Palm forest, marshland, and mangrove) that occur throughout the state of Maranhão, in northeastern Brazil. We sampled 90 sites (45 preserved and 45 anthropogenic areas) where we collected 26,036 specimens: 15,023 calliphorids (11 species), 231 mesembrinellids (one species), and 10,772 sarcophagids (52 species). Four environmental factors, canopy openness, temperature, leaf litter depth, and vegetation height, contributed most to the separation of preserved and anthropogenic sites in all five phytophysiognomies. Leaf litter depth was positively associated with the species richness of the calliphorids and mesembrinellids (C+ M group), while tree/shrub density, vegetation height, and temperature were associated negatively with the richness of the sarcophagids. Tree/shrub density and vegetation height were also associated negatively with abundance in both C+M and sarcophagid species. Overall, then, the structural characteristics of the environment affected the species richness and abundance, and deforestation may favor certain synanthropic species, leading to a decrease in the richness and abundance of the species that are adapted to preserved environments.

Comparative analysis of the mitochondrial genomes of flesh flies and their evolutionary implication.

Flesh flies (Diptera: Sarcophagidae) include a large and widely distributed rapid radiation within the Calyptratae. They are vital for the ecosystem, as well as economic, forensic, and evolutionary studies, because of their extremely diverse habits as larvae. Phylogenetic studies of Sarcophagidae have been reaching convergence, which leads the opportunity to elucidate the evolution of these fast-evolving insects from the perspective of mitochondrial genome. Complete mitochondrial genomes of eight species were sequenced, and comparative mitochondrial genomic analysis between subfamilies were conducted. Mitochondrial genomes of these flesh flies are conserved in gene content with gene arrangement, same as the inferred ancestral insect, and the nucleotide composition is highly biased towards A + T like other flesh flies. The evolutionary rates of Sarcophagidae vary considerably across subfamilies, with that of Miltogramminae higher than the other two subfamilies. Phylogenetic analysis strongly supports monophyly of Sarcophagidae and each subfamily, with subfamily-level relationship inferred as (Sarcophaginae, (Miltogramminae, Paramacronychiinae)). The main topological inconsistency of all reconstructions is the relationship within Miltogramminae and Sarcophaga, which might be caused by their rapid evolution. Our study indicates that the mitochondrial genomes of flesh flies are highly conserved, and they are practically useful for phylogenetic inference of calyptrates.

Description of Two New Species of Dexosarcophaga (Diptera: Sarcophagidae) From the Coastal Environments of the Brazilian Amazon.

Two new species of Dexosarcophaga Townsend (Diptera: Sarcophagidae) are described, both from the coastal zone of the Brazilian Amazon: Dexosarcophaga salgada sp. nov. and Dexosarcophaga klycyae sp. nov.

New carrion-visiting flesh flies (Diptera: Sarcophagidae) from tropical dry forests of Colombia and their phylogenetic affinities.

Many flesh flies of the subfamily Sarcophaginae are very competitive carrion visitors and breeders, and several are synanthropic species of medical and forensic importance. The knowledge of these taxa is still limited, which is more noticeable in sub-sampled ecosystems such as the tropical dry forests of Colombia. Four new species, Blaesoxipha (Gigantotheca) wajiira sp. nov., Lepidodexia deborarangoa sp. nov., Oxysarcodexia luriza sp. nov. and Udamopyga iku sp. nov., are described from tropical dry forests in the Colombian departments of Atlántico, Bolívar, Cesar and La Guajira, all collected using Van Someren-Rydon traps baited separately with fermented fruit, decomposing fish and human feces. The species B. (G.) wajiira sp. nov. is probably synanthropic, while the remaining three new species avoid habitats in and around built-up areas. A phylogenetic parsimony analysis based on 222 morphological characters across 121 taxa of flesh flies recovers each new species nested within their respective genus. The subgeneric affiliation of L. deborarangoa sp. nov. within the large and diverse genus Lepidodexia (sensu lato) is inconclusive, and potential issues of relying solely on external morphological characters for generic and subgeneric classifications are highlighted. The species Lepidodexia (Notochaeta) woodi (Lopes), Oxyvinia wicharti (Lopes) and Sarcofahrtiopsis spinetta Mulieri & Dufek are new records for Colombia. COI sequence data are provided for U. iku sp. nov., L. (N.) woodi and O. wicharti.

Evaluation of environmental and local factors on urban sarcosaprophagous Diptera: a case study with Sarcophagidae.

The urbanization process affects the structure of insect communities, favouring biotic homogenization processes by promoting adequate conditions for the establishment of synanthropic species. Considering the scarce knowledge about the ecology of Neotropical sarcophagids, we compared the flesh fly community from two different urban zones in two contrasting seasons and examined the response of the Sarcophagidae assemblage to landscape and microhabitat variables. Adult flies were collected in 13 sites with two traps (total number of traps 26) baited with 200 g of hydrated bone meal. A total of 2312 adult sarcophagids were collected belonging to 27 species; Oxysarcodexia paulistanensis (Mattos, 1919) represented >48% of the total sample. The local factor of insolation explained both the abundance and species richness of sarcophagids, collections being higher and richer in traps exposed to sunlight compared with traps placed in the shade regardless of the season. Richness was also affected by the environment, being higher in traps exposed in suburban rather than urban sites. The combination of these factors (sub-urban environment and availability of open areas) could explain the Sarcophagidae assemblage. The average number of sarcophagid species collected was related to the season, providing a good opportunity to assess the temporal variability of this family in the region.

Comparative analysis of mitochondrial genomes among the subfamily Sarcophaginae (Diptera: Sarcophagidae) and phylogenetic implications.

The subfamily Sarcophaginae is extremely diverse in morphology, habit and geographical distribution, and usually considered to be of significant ecological, medical, and forensic significance. In the present study, 18 mitochondrial genomes (mitogenomes) of sarcophagid flies were first obtained. The rearrangement and orientation of genes were identical with that of ancestral insects. The degrees of compositional heterogeneity in the datasets were extremely low. Furthermore, 13 protein-coding genes were evolving under purifying selection. The phylogenic relationship of the genus-group taxa Boettcheria + (Sarcophaga + (Peckia + (Ravinia + Oxysarcodexia))) was strongly supported. Four subgenera were recovered as monophyletic (Liopygia, Liosarcophaga, Pierretia, Heteronychia) in addition to Parasarcophaga as polyphyletic. The sister-relationships between S. dux and S. aegyptiaca, S. pingi and S. kawayuensis were recovered, respectively. Moreover, the molecular phylogenetic relationships among the subgenera Helicophagella, Kozlovea, Kramerea, Pandelleisca, Phallocheira, Pseudothyrsocnema, Sinonipponia and Seniorwhitea were rarely put forward prior to this study. This study provides insight into the population genetics, molecular biology, and phylogeny for the subfamily Sarcophaginae, especially for the subgeneric classification of Sarcophaga. However, compared with the enormous species diversity of flesh flies, the available mitogenomes are still limited for recovering the phylogeny of Sarcophaginae.

Flesh Flies (Diptera: Sarcophagidae) Attracted to a Snake Carcass (Boa constrictor) in Yucatan Peninsula, Mexico.

A study was carried out to determine Sarcophagidae diversity attracted to the different stages of decomposition of a Boa constrictor cadaver during late winter in the Yucatan Peninsula. As a result of this study, seven species of Sarcophagidae were documented, Oxysarcodexia conclausa (Walker, 1861) (Diptera: Sarcophagidae), Peckia (Euboettcheria) volucris (Wulp, 1895) (Diptera: Sarcophagidae), Blaesoxipha (Gigantotheca) plinthopyga (Wiedemann, 1830) (Diptera: Sarcophagidae), Oxysarcodexia amorosa (Schiner, 1868) (Diptera: Sarcophagidae), Ravinia derelicta (Walker, 1853) (Diptera: Sarcophagidae), Ravinia effrenata (Walker, 1861) (Diptera: Sarcophagidae), and Titanogrypa (Cucullomyia) placida (Aldrich, 1925) (Diptera: Sarcophagidae), The last five species listed are the first documentation of their presence in the Yucatan Peninsula in Mexico. Sarcophagids were present throughout the decomposition process; however, the greatest abundance and diversity of this family were present during the advanced decay stage. This is the first work on flesh flies in the region and the first in the country that has focused on wildlife.

Two new records of flies causing myiasis from Saudi Arabia with a survey of flies parasitizing goats and sheep in Jazan Region.

Despite the medical, veterinary and forensic relevance of myiasis-causing flies, knowledge of their diversity in Saudi Arabia is limited especially in the southern region. Therefore, a survey of myiasis-causing flies in the Jazan region was carried out using Red Top Fly Catcher traps baited with either decomposing beef liver or a lure composed primarily from fishmeal during the period April 2013-March 2014. Twelve known species were identified and recorded in this study, seven species of them belonging to Calliphoridae (Chrysomya, Lucilia, and Hemipyrella) and five species belonging to Sarcophagidae (Sarcophaga). Two of these species were recorded for the first time for Saudi Arabia, namely Hemipyrella pulchra (Wiedemann, 1830) and Sarcophaga (Liosarcophaga) exuberans Pandellé, 1896. Images of the species recorded are also provided for the first time. The results expand the knowledge of geographical distribution, fauna, and habitat of the myiasis-causing flies in Saudi Arabia. Biological information and world-wide geographical distribution of these species are included together with some taxonomic remarks.

Genome and Ontogenetic-Based Transcriptomic Analyses of the Flesh Fly, Sarcophaga bullata.

The flesh fly, Sarcophaga bullata, is a widely-used model for examining the physiology of insect diapause, development, stress tolerance, neurobiology, and host-parasitoid interactions. Flies in this taxon are implicated in myiasis (larval infection of vertebrates) and feed on carrion, aspects that are important in forensic studies. Here we present the genome of S. bullata, along with developmental- and reproduction-based RNA-Seq analyses. We predict 15,768 protein coding genes, identify orthology in relation to closely related flies, and establish sex and developmental-specific gene sets based on our RNA-Seq analyses. Genomic sequences, predicted genes, and sequencing data sets have been deposited at the National Center for Biotechnology Information. Our results provide groundwork for genomic studies that will expand the flesh fly's utility as a model system.

Comparative Mitogenomic Analysis of Forensically Important Sarcophagid Flies (Diptera: Sarcophagidae) and Implications of Species Identification.

The flesh flies (Diptera: Sarcophagidae) are significant in forensic investigations. The mitochondrial genome (mitogeome) has been widely used as genetic markers for phylogenetic analysis and species identification. To further understand the mitogenome-level features in Sarcophagidae, the complete mitogenome of Sarcophaga formosensis (Kirneret Lopes, 1961) (Diptera: Sarcophagidae) and Sarcophaga misera (Walker, 1849) (Diptera: Sarcophagidae) was firstly sequenced, annotated, and compared with other 13 Sarcophagidae species. The result indicated that the gene arrangement, gene content, base composition, and codon usage were conserved in the ancestral arthropod. Evolutionary rate of the mitogenome fragments revealed that the nonsynonymous and synonymous substitution rates (Ka and Ks) ratio was less than 1.00, indicating these variable sites under strong purifying selection. Almost all transfer RNA genes (tRNAs) have typical clover-leaf structures within these sarcophagid mitogenomes, except tRNA-Ser (AGN) is lack of the dihydrouridine arm. This comparative mitogenomic analysis sheds light on the architecture and evolution of mitogenomes in the Sarcophagidae. Phylogenetic analyses containing the interspecific distances from different regions in these species provided us new insights into the application of these effective genetic markers for species identification of flesh flies.

Wing morphometric analysis of forensically important flesh flies (Diptera: Sarcophagidae) in Thailand.

Flesh flies are insects of forensic importance as the larvae associated with human remains can be used to estimate a minimum post-mortem interval (PMImin) in most cases. And, because life-history traits can vary across species, correct identification is a mandatory step before being used as evidence. Adult flesh flies are extremely similar in general appearance, which causes difficulty in species identification as it is largely based on the morphology of the male genitalia; this also makes it difficult to identify females. Currently, landmark-based geometric morphometric analysis of insect wings has proven to be a valuable tool for species identification. Herein, we applied wing morphometric analysis of 524 flesh fly specimens comprising 12 species from Thailand. The right wing of each specimen was removed, mounted on a microscope slide, photographed, and digitized using 18 landmarks. Wing shape variation among genera and species were analyzed using canonical variate analysis, while wing shape variation between sexes of each species was analyzed using discriminant function analysis. A cross-validation test was used to evaluate the reliability of classification. Results of this study demonstrate wing shape can be used to separate genera and species, and distinguish between sexes of the same species, with high reliability. Therefore, the landmark-based geometric morphometric analysis of wings is a useful additional method for species and sex discrimination of flesh flies.

A novel use of infra-red spectroscopy (NIRS and ATR-FTIR) coupled with variable selection algorithms for the identification of insect species (Diptera: Sarcophagidae) of medico-legal relevance.

Unequivocal identification of fly specimens is an essential requirement in forensic entomology. Herein, a simple, non-destructive and rapid method based on two vibrational spectroscopy techniques [Near-Infrared Spectroscopy (NIRS) and attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy] coupled with variable selection techniques such as genetic algorithm-linear discriminant analysis (GA-LDA) and successive projection algorithm-linear discriminant analysis (SPA-LDA) were applied for identifying and discriminating six species of flesh flies (Diptera: Sarcophagidae) native to Neotropical regions. This novel approach is based on the unique spectral "fingerprints" of their biochemical composition. One hundred sixty (160) NIRS and FT-IR specimens (120 male, 40 female) were acquired; different pre-processing methods such as baseline correction, derivative and Savitzky-Golay smoothing were also performed. In addition, the multivariate classification accuracy results were tested based on sensitivity, specificity, positive (or precision) and negative predictive values, Youden index, positive and negative likelihood ratios. Principal components analysis (PCA) was employed for male vs. female category using NIRS, strongly showing the separation between the classes with only three principal components and 99% explained variance. Differentiation between the genera Oxysarcodexia, Peckia and Ravinia was efficiently confirmed by both techniques. In comparison with other biological methods, this approach represents an effective choice for fast and non-destructive identification in forensic entomology.

DNA barcoding for identifying synanthropic flesh flies (Diptera, Sarcophagidae) of Colombia.

The first step for a successful use of any insect as indicator in forensic sciences is providing a precise taxonomic identification at species level. Due to morphology-based identification of Sarcophaginae flies (Diptera, Sarcophagidae) is often difficult and requires strong taxonomic expertise, their use as forensic indicators has been limited. Consequently, molecular-based approaches have been accepted as alternative means of identification. Thus, we aimed testing the efficiency of the barcode region of the mitochondrial cytochrome oxidase subunit I (COI) gene for identification of synanthropic flesh flies of several species of the genera Peckia, Oxysarcodexia, Ravinia, and Tricharaea collected in Colombia. The 645-bp fragment of COI was amplified and aligned (215 parsimoniously informative variable sites). We calculated Kimura two-parameter genetic distances and reconstruct a Neighbor-Joining phylogenetic tree. Our Neighbor-Joining tree recovered all species as monophyletic, and confirmed a new species of the genus Ravinia as also indicated by the interspecific genetic divergences and morphological observations. We obtained a 100% of identification success. Thus, the COI barcodes showed efficiency as an alternative mean of identification of species of flesh flies collected on decaying organic matter in Colombia.

A new flesh fly species (Diptera: Sarcophagidae) parasitic on leptodactylid frogs.

Here we describe a new species of sarcophagid fly reared from larvae found feeding on three species of leptodactylid frogs collected in subtropical Corrientes Province in northeastern Argentina. Our species description is based on adult male and female external morphology and genitalia. Adult males and females of the new species were associated with certainty because all specimens studied were reared from single clutches of larvae on each of three infested frogs. Thus, adult males and females reared from each clutch were siblings that emerged almost simultaneously. The paper provides line drawings, high-resolution photographs, and SEM images to aid in identifying Lepidodexia (Notochaeta) adelina sp. nov. and to distinguish the new species from its closely related congeners. We include brief notes on the host species and on larval feeding behavior of L. (N.) adelina. This is the first and only species in the genus Lepidodexia reported to parasitize leptodactylid frogs. Further, our observations reported here are the first to fully document a completed life cycle by sarcophagid fly larvae on three different species of leptodactylid frogs and the only well-documented case of myiasis of an amphibian in Argentina.

Insects Associated with Pig Carrion in Two Environments of the Brazilian Savanna.

Studies of carrion fauna have increased in Brazil and have contributed to the knowledge of this fauna in the national territory. Brazil has continental dimensions and presents various biomes in its territorial area. Most of the carrion fauna have seldom been studied, and this is especially true for the Savanna or "Cerrado." The present research examined the fauna visiting carrion in two environments and two seasons in a rural area situated in a region of Cerrado in southeastern Brazil. The two environments studied were a pasture and a fragment of semi-deciduous forest. Samples were collected during the dry and humid seasons of the year. The study identified the diversity and relative abundance of species attracted to pig carcasses (Sus scrofa L.) exposed in each environment and season. Eight pig carcasses (10 ± 1 kg) were placed in traps, and adult insects attracted to the traps were collected during the decay of the carcasses. A total of 92,489 insects were collected during the experiments. The Diptera was the most frequent order (92.2%) represented by 27 families and at least 124 species. This was followed by Coleoptera (4.4%) represented by 14 families and at least 65 species. Sarcophagidae had the greatest diversity of species, followed by Muscidae and Calliphoridae. Several species were collected only during a specific period of the year or in a single environment. The results are important from an ecological point of view and for Medico-Legal Forensic Entomology.

Molecular phylogeny of Miltogramminae (Diptera: Sarcophagidae): Implications for classification, systematics and evolution of larval feeding strategies.

Miltogramminae is one of the phylogenetically most poorly studied taxa of the species-rich family Sarcophagidae (Diptera). Most species are kleptoparasites in nests of solitary aculeate wasps and bees, although parasitoids and saprophagous species are also known, and the ancestral miltogrammine life habit remains unsettled. Here, we present for the first time a comprehensive phylogenetic tree consisting of 58 representatives of Miltogramminae, reconstructed using sequence data from three mitochondrial (COI, cytB, ND4) and one nuclear (Ef-1α) genes. Our phylogenetic hypothesis suggests that: (1) Miltogramminae are sister to Paramacronychiinae, (2) Miltogramminae can be divided into the "lower miltogrammines" containing two clades of mainly saprophages and a clade of "higher miltogrammines" with mainly kleptoparasitic species, (3) only three genera turn out to be non-monophyletic: Miltogramma, Senotainia and Pterella and (4) the genus Sarcotachina, which traditionally has been considered as belonging to the Paramacronychiinae, is placed in one of the clades of "lower miltogrammines". Ancestral state reconstruction of larval feeding strategy and five larval characters reveals that the ancestor of Miltogramminae was likely a saprophage retaining plesiomorphic oral ridges and a cephaloskeleton with sclerotized dorsal bridge. Synapomorphies like large pseudocephalic sensory organs and well-developed cuticular sculpture suggest that the ancestral first instar larva actively searched for a buried food supply.