Autogeny and endocrine involvement in the reproduction of tropical flesh flies (Diptera: Sarcophagidae)

Autogeny is highly developed among the Sarcophagidae in tropical Africa. Unlike their temperate region relatives, the tropical species examined can mature the first batch of eggs without an adult protein meal, and they do so without the long period of delay usually seen in autogenous species. Protein deprivation, however, does reduce the number of eggs matured by about one-third. The tropical and temperate flies appear to share a common endocrine mechanism controlling egg maturation. Removal of the medial neurosecretory cells of the pars intercerebralis prevents egg maturation. Reimplantation of clusters of MNC restores the ability to mature eggs. Extirpation of the corpora allata reduces the percentage of females that mature eggs to about 70%, and the females that do mature eggs produce fewer eggs. The significant difference between the tropical and temperate species centers on the trigger that sets the neuroendocrine system into action. For the tropical species, the brain is activated immediately and does not require the trigger of a protein meal.     

Anchored hybrid enrichment challenges the traditional classification of flesh flies (Diptera: Sarcophagidae)

Sarcophagidae is one of the most species-rich families within the superfamily Oestroidea. This diversity is usually represented by three lineages: Miltogramminae, Paramacronychiinae and Sarcophaginae. Historically, the phylogenetic relationships among these lineages have been elusive, due to poorly supported hypotheses or small taxon sets, or both. This study provides a dramatic increase in molecular data, more balanced sampling of all three lineages from all biogeographical regions and a reassessment of morphological characters using scanning electron microscopy in the most comprehensive assessment of subfamily-level phylogeny in Sarcophagidae to date. This analysis of the largest molecular dataset ever produced for a phylogenetic analysis of a fly lineage, with 950 loci from anchored hybrid enrichment comprising 435 930 bp from 101 species, revealed Paramacronychiinae as sister to Miltogramminae, not to Sarcophaginae, as suggested by adult morphology. Maximum likelihood analysis produced a well-supported topology, with 91% of the nodes receiving strong bootstrap proportions (> 97%). In contrast to the molecular data, three out of nine morphological characters studied point to a sister-group relationship of (Sarcophaginae + Paramacronychiinae) and the remaining six characters are either silent on subfamily relationships or in need of further study. Re-examination of morphological structures provides new insights into the evolution of male genitalic traits within Sarcophagidae and highlights their convergence producing conflicting phylogenetic signal. Our phylogeny reconciles older and widely used systems of classification with tree-based thinking and sets up a classification of flesh flies that is more aligned with their evolutionary history.     

A large‐scale molecular phylogeny of flesh flies (Diptera: Sarcophagidae)

The available data for Sarcophagidae in GenBank were analysed in order to reconstruct the most comprehensive phylogeny to date. GenBank was explored for nine markers that are commonly used in various molecular and phylogenetic studies of flesh flies. We obtained data for 187 species and constructed an aligned dataset with 9241 characters. However, the matrix suffered from 74% missing data due to a low number of sequences for some markers and in most of the cases only short fragments of the analysed genes were available. The reconstructed tree was taxonomically biased towards the subfamilies Paramacronychiinae (12% of the described species) and Sarcophaginae (8.6% of the described species) and specifically the genus Sarcophaga. The third subfamily Miltogramminae was represented by only 0.7% of described species. Moreover, about half of the included species were of forensic importance, while the percentage of such species in the entire family was estimated at 7%. Many nodes had very low support, so in order to increase the support and thereby identify a ‘core topology’, we pruned ‘rogue’ taxa and applied different substitution models. Both strategies improved support considerably, although some nodes still were left unresolved. An analysis of the distribution of bootstrap values across chronograms showed that the weakest phylogenetic signal is restricted to that part of the tree which coincides with the onset of rapid radiations mainly within the genus Sarcophaga. Our study is concordant with phylogenies obtained by other authors, with the most noteworthy exception being the subfamily Paramacronychiinae emerging as paraphyletic with regard to the Miltogramminae, which is in strong conflict with morphological evidence. We discuss the new findings in the light of traditional taxonomical classifications of Sarcophagidae and recent molecular studies.     

Phylogenetic relationships of flesh flies in the subfamily Sarcophaginae based on three mtDNA fragments (Diptera: Sarcophagidae)

In an effort to improve our knowledge of the phylogenetic relationships among species and genera of the subfamily Sarcophaginae, we analysed data from three mitochondrial gene fragments. Sequence data for portions of the genes cytochrome oxidase I (COI), cytochrome oxidase II (COII) and dehydrogenase subunit 4 (ND4) were obtained from 43 species of Sarcophagidae representing 15 genera. We used a Bayesian approach to simultaneously choose how best to partition the data and which substitution model to apply to each partition. Phylogenetic relationships were inferred using Bayesian Inference and Maximum Likelihood methods. Our results are consistent with monophyly of the subfamily Sarcophaginae (posterior probability 1; bootstrap support 93%), as well as with monophyly of several genera within the Sarcophaginae (including Sarcophaga s.l.; posterior probability 1; bootstrap support 97%). We found support for a sister‐group relationship between Ravinia Robineau‐Desvoidy and Oxysarcodexia Townsend, which has been hypothesised by past authors on the basis of morphological similarities, although this was supported only in the Bayesian analyses (posterior probability 0. 81–0. 98), and for some novel supra‐generic clades. Contrary to a recent morphological hypothesis, we do not find Helicobia Coquillett to be nested within Sarcophaga Meigen; our data suggest, but do not strongly support, a hypothesis that Peckia Robineau‐Desvoidy is the sister group to Sarcophaga.     

Three new species of Sarcophaga Meigen found during ecological studies on flesh flies (Diptera: Sarcophagidae) in Thailand

Three new species of Sarcophaga Meigen, 1826 are described from Thailand: Sarcophaga ( Sarcosolomonia ) circa sp. nov., S . ( Mehria ) lanna sp. nov. and S . ( Rosellea ) suthep sp. nov.     

A comparison of the responses of tropical and temperate flies (Diptera: Sarcophagidae) to cold and heat stress

Summary Two flesh fly species from the tropical lowlands (Peckia abnormis and Sarcodexia sternodontis) were more susceptible to both cold-shock and heatshock injury than temperate flies (Sarcophaga crassipalpis and S. bullata) and a fly from a tropical high altitude (Blaesoxipha plinthopyga). A brief (2-h) exposure to 0°C elicits a protective response against subsequent cold injury at–10°C in the temperate flies and in B. plinthopyga but no such response was found in the flies from the tropical lowlands. However, both tropical and temperate flies could be protected against heat injury (45°C) by first exposing them to a mild heat shock (2 h at 40°C). The supercooling point is not a good indicator of cold tolerance: supercooling points of pupae were similar in all species, ranging from–18.9 to–23.0°C, and no differences were found between the tropical and temperate species. Among the temperate species, glycerol, the major cryoprotectant, can be elevated by short-term exposure to 0°C, but glycerol could not be detected in the tropical flies. Low-temperature (0°C) exposure also increased hemolymph osmolality of the temperate species, but no such increase was observed in the tropical lowland species. Adaptations to temperature stress thus differ in tropical and temperate flesh flies: while flies from both geographic areas share a mechanism for rapidly increasing heat tolerance, only the temperate flies appear capable of responding rapidly to cold stress. The presence of a heat shock response in species that lack the ability to rapidly respond to cold stress indicates that the biochemical and physiological bases for these two responses are likely to differ.     

Malate dehydrogenase isozymes in five species of sarcophagid flies (Sarcophagidae: Diptera)

Malate dehydrogenase banding patterns were analyzed by polyacrylamide gel electrophoresis in Sarcophaga ruficornis, S. argyrostoma, S. dux, S. invaria and S. peregrina. Two distinct zones of MDH activity, attributed to be the product of two gene loci, namely, MDH-1 and MDH-2 were observed. The MDH-1 locus revealed polymorphism for three electrophoretic phenotypes which are governed by two electromorphs MDH-1^a and MDH-1^b. The MDH banding pattern does not reveal any notable interspecific difference.     

Sarcophaga (Liosarcophaga) dux (Diptera: Sarcophagidae): A flesh fly species of medical importance

Background

Although tropical climate of Thailand is suitably endowed with biodiversity of insects, flies of medical importance is not well investigated. Using information from literature search, fly survey approach and specialist’s experience, we review database of Sarcophaga (Liosarcophaga) dux Thomson (Diptera: Sarcophagidae), one of the priorities flesh fly species of medical importance in Thailand.

Results

This review deals with morphology, bionomics and medical involvement. Important morphological characteristics of egg, larva, puparia and adult were highlighted with illustration and/or micrographs. Search pertaining to molecular analysis used for fly identification and developmental rate of larvae were included. Medical involvement of larvae was not only myiasis-producing agent in humans and animals, but associated with human death investigations.

Conclusions

This information will enable us to accurate identify this species and to emphasis the increase medically important scene in Thailand.     

The complete mitochondrial genome of the flesh fly, Sarcophaga impatiens Walker (Diptera: Sarcophagidae)

Approximately 2500 fly species comprise the Sarcophagidae family worldwide. The complete mitochondrial genome of the carrion-breeding, forensically important Sarcophaga impatiens Walker (Diptera: Sarcophagidae) from Australia was sequenced. The 15,169 bp circular genome contains the 37 genes found in a typical Metazoan genome: 13 protein-coding genes, 2 ribosomal RNA genes and 22 transfer RNA genes. It also contains one non-coding A t T-rich region. The arrangement of the genes was the same as that found in the ancestral insect. All the protein initiation codons are ATN, except for cox1 that begins with TCG (encoding S). The 22 tRNA anticodons of S. impatiens are consistent with those observed in Drosophila yakuba, and all form the typical cloverleaf structure, except for tRNA-Ser((AGN)) that lacks the DHU arm. The mitochondrial genome of Sarcophaga presented will be valuable for resolving phylogenetic relationships within the family Sarcophagidae and the order Diptera, and could be used to identify favourable genetic markers for species identifications for forensic purposes.     

Towards a phylogeny of the flesh flies (Diptera: Sarcophagidae): morphology and phylogenetic implications of the acrophallus in the subfamily Sarcophaginae

The morphology of the acrophallus, the distal portion of the male phallus carrying the phallotreme, was studied in 72 exemplar species representing 56 genera and subgenera of the flesh fly subfamily Sarcophaginae. For 42 of those species, scanning electron microscopy was used to clarify the phallic morphology. Terms used to describe the male genitalia were updated based on new interpretations of homology. Male genitalic characters, combined with other morphological characters of adult males and females and of larvae, were used to construct a phylogeny. The monophyly of the subfamily was supported, and some generic‐level sister‐group relationships proposed in the literature, but without previous cladistic analyses, were also supported. The genus Blaesoxipha Loew, as currently recognized, was not monophyletic in our analysis. The genus Helicobia Coquillett is synonymized with Sarcophaga Meigen syn. nov. and treated as a subgenus of the latter. The Sarcophaga subgenera Neobellieria Blanchard and Mehria Enderlein were not monophyletic. Many of the clades in the analysis were supported primarily or exclusively by male genitalic character states, highlighting the importance of the male genitalia as a source of morphological characters for sarcophagine phylogeny. © 2010 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010, 158 , 740–778.     

Vitamin requirements of the cultured flesh fly cells, Sarcophaga peregrina (Diptera, Sarcophagidae)

Essential vitamins for the growth of a cell line derived from the flesh fly, Sarcophaga peregrina, were determined. By examining the survivability of continuous passages of the cells in the chemically defined medium lacking one vitamin, thiamine, riboflavin, pantothenate and either niacin or niacinamide were found to be essential for the continuous growth of the flesh fly cells in vitro. [Originally published in Volume 37, Archives of Insect Biochemistry and Physiology, 37:283-286 (1998).] Copyright 1998 Wiley-Liss, Inc.     

Vitamin requirements of the cultured flesh fly cells,Sarcophaga peregrina (Diptera,Sarcophagidae)

Essential vitamins for the growth of a cell line derived from the flesh fly, Sarcophaga peregrina, were determined. By examining the survivability of continuous passages of the cells in the chemically defined medium lacking one vitamin, thiamine, riboflavin, pantothenate and either niacin or niacinamide were found to be essential for the continuous growth of the flesh fly cells in vitro. Arch. Insect Biochem. Physiol. 37:283–286, 1998. © 1998 Wiley-Liss, Inc.     

The Sarcophagidae (Diptera) of the Middle East

A list of 285 species of Sarcophagidaе in the Middle East countries is presented with distributional data, including Bahrain (3 species), Cyprus (46), Egypt (both African and Asian parts) (114), Iran (83), Iraq (17), Israel (113), Jordan (14), Kuwait (3), Lebanon (13), Oman (2), Gaza Strip (5), Palestinian Authority (42), Quatar (1), Saudi Arabia (37), Syria (42), Turkey (both European and Asian parts) (157), United Arab Emirates (14) and Yemen (15). Three new synonyms are established: Blaesoxipha delilah Lehrer, 2006 = Agriella setosa Salem, 1938, syn. n.; Blaesoxipha nahaliana Lehrer, 2008 = Blaesoxipha popovi Rohdendorf, 1937, syn. n.; and Liosarcophaga daccanella Lehrer, 2008 = Liosarcophaga (s. str.) dux (Thomson, 1869), syn. n. Four new combinations for species names are proposed: Liopygia (Engelisca) adhamae (Lehrer & Abou-Ziad, 2008), comb. n.; Liosarcophaga (s. str.) pedestris (Villeneuve, 1910), comb. n.; Liosarcophaga (Pandelleisca) theodori (Lehrer, 1998), comb. n., and Liosarcophaga (Pharaonops) tewfiki (Salem, 1940), comb. n.     

DNA barcoding of tropical black flies (Diptera: Simuliidae) of Thailand

The ecological and medical importance of black flies drives the need for rapid and reliable identification of these minute, structurally uniform insects. We assessed the efficiency of DNA barcoding for species identification of tropical black flies. A total of 351 cytochrome c oxidase subunit 1 sequences were obtained from 41 species in six subgenera of the genus Simulium in Thailand. Despite high intraspecific genetic divergence (mean = 2.00%, maximum = 9.27%), DNA barcodes provided 96% correct identification. Barcodes also differentiated cytoforms of selected species complexes, albeit with varying levels of success. Perfect differentiation was achieved for two cytoforms of Simulium feuerborni, and 91% correct identification was obtained for the Simulium angulistylum complex. Low success (33%), however, was obtained for the Simulium siamense complex. The differential efficiency of DNA barcodes to discriminate cytoforms was attributed to different levels of genetic structure and demographic histories of the taxa. DNA barcode trees were largely congruent with phylogenies based on previous molecular, chromosomal and morphological analyses, but revealed inconsistencies that will require further evaluation.     

Diapausing pupae of the flesh fly Sarcophaga crassipalpis (Diptera: Sarcophagidae) are more resistant to inoculative freezing than non-diapausing pupae

The resistance of diapausing (overwintering) and non‐diapausing (summer) Sarcophaga crassipalpis (Diptera: Sarcophagidae) pupae to inoculative freezing was examined. Although both types of pupae resisted inoculative freezing after 24‐h submergence in water, diapausing pupae were overall significantly more resistant than non‐diapausing pupae. Exposing the thin pupal cuticle by removing the ends of the puparial case eliminated the capacity of both pupal types to resist inoculative freezing, indicating that resistance to inoculative freezing resides with the puparium. Pupae submerged in surfactant solution were significantly less resistant to inoculative freezing than those submerged in water. Furthermore, the puparial water content of pupae submerged in surfactant solution was significantly greater than that of puparia from pupae submerged in water. Surfactant may have promoted inoculative freezing by facilitating the spread of water over the surface of and into the puparium, thereby creating bridges between external ice and pupal body fluids. Extracting puparial surface lipids with chloroform/methanol (2 : 1, v:v) decreased the resistance of non‐diapausing pupae to inoculative freezing but did not significantly affect that of diapausing pupae. This finding indicates that the puparium of diapausing pupae contains protection against inoculative freezing separate from its surface lipids. This barrier may be important in protecting the freezing‐intolerant overwintering pupae against inoculative freezing within their soil hibernaculum.     

Mitochondrial DNA sequence-based phylogenetic relationship among flesh flies of the genus <Emphasis Type="Italic">Sarcophaga</Emphasis> (Sarcophagidae: Diptera)

The phylogenetic relationships among flesh flies of the family Sarcophagidae has been based mainly on the morphology of male genitalia. However, the male genitalic character-based relationships are far from satisfactory. Therefore, in the present study mitochondrial DNA has been used as marker to unravel genetic relatedness and to construct phylogeny among five sympatric species of the genus Sarcophaga. Two mitochondrial genes viz., cytochrome oxidase subunit 1 (COI) and NAD dehydrogenase subunit 5 (ND5) were sequenced and genetic distance values were calculated on the basis of sequence differences in both the mitochondrial genes. The data revealed very few genetic difference among the five species for the COI and ND5 gene sequences.     

Isolation and identification of a cAMP generating peptide from the flesh fly,Neobellieria bullata (Diptera: Sarcophagidae)

The Manduca sexta Malpighian tubule assay system, developed to monitor adenylate cyclase activity, was used in combination with HPLC to isolate a novel cAMP generating peptide from 350,000 whole flesh flies, Neobellieria bullata. Mass spectrometry revealed a molecular mass of 5,047 daltons, and Edman degradation the following sequence: AGAEAEKLSGLSKYFNGTTMAGRANVAKATYAVIGLIIAYNVMKPKKK. This 48-mer peptide, called Neb-cGP, does not belong to the corticotropin releasing factor family of insect diuretic peptides. Electrophoresis and subsequent immunoblotting of peptides immunoprecipitated from a homogenate of entire flies showed that one fly contained approximately 0.003 to 0.03 μg Neb-cGP and that 10 μg represents the lowest immunostainable amount on a Western blot. © 1996 Wiley-Liss, Inc.     

The ectoparasitic wasp Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae) differentially affects cells mediating the immune response of its flesh fly host,Sarcophaga bullata Parker (Diptera: Sarcophagidae)

In this study, we examined cellular immune responses in the flesh fly, Sarcophaga bullata, when parasitized by the ectoparasitoid Nasonia vitripennis. In unparasitized, young pharate adults and third instar, wandering larvae of S. bullata, four main hemocyte types were identified by light microscopy: plasmatocytes, granular cells, oenocytoids, and pro-hemocytes. Parasitism of young pharate adults had a differential effect on host hemocytes; oenocytoids and pro-hemocytes appeared to be unaltered by parasitism, whereas adhesion and spreading behavior were completely inhibited in plasmatocytes and granular cells by 60 min after oviposition. The suppression of spreading behavior in granular cells lasted the duration of parasitism. Plasmatocytes were found to decline significantly during the first hour after parasitism and this drop was attributed to cell death. Melanization and clotting of host hemolymph did not occur in parasitized flies, or the onset of both events was retarded by several hours in comparison to unparasitized pharate adults. Hemocytes from envenomated flies were altered in nearly identical fashion to that observed for natural parasitism; the total number of circulating hemocytes declined sharply by 60 min post-envenomation, the number of plasmatocytes declined but not granular cells, and the ability of plasmatocytes and granular cells to spread when cultured in vitro was abolished within 1 h. As with parasitized hosts, the decrease in plasmatocytes was due to cell death, and inhibition of spreading lasted until the host died. Isolated crude venom also blocked adhesion and spreading of these hemocyte types in vitro. Thus, it appears that maternally derived venom disrupts host immune responses almost immediately following oviposition and the inhibition is permanent. The possibility that this ectoparasite disables host defenses to afford protection to feeding larvae and adult females is discussed.