The Acquisition and Retention of Lumpy Skin Disease Virus by Blood-Feeding Insects Is Influenced by the Source of Virus, the Insect Body Part, and the Time since Feeding.

Lumpy skin disease virus (LSDV) is a poxvirus that causes severe systemic disease in cattle and is spread by mechanical arthropod-borne transmission. This study quantified the acquisition and retention of LSDV by four species of Diptera (Stomoxys calcitrans, Aedes aegypti, Culex quinquefasciatus, and Culicoides nubeculosus) from cutaneous lesions, normal skin, and blood from a clinically affected animal. The acquisition and retention of LSDV by Ae. aegypti from an artificial membrane feeding system was also examined. Mathematical models of the data were generated to identify the parameters which influence insect acquisition and retention of LSDV. For all four insect species, the probability of acquiring LSDV was substantially greater when feeding on a lesion compared with feeding on normal skin or blood from a clinically affected animal. After feeding on a skin lesion LSDV was retained on the proboscis for a similar length of time (around 9 days) for all four species and for a shorter time in the rest of the body, ranging from 2.2 to 6.4 days. Acquisition and retention of LSDV by Ae. aegypti after feeding on an artificial membrane feeding system that contained a high titer of LSDV was comparable to feeding on a skin lesion on a clinically affected animal, supporting the use of this laboratory model as a replacement for some animal studies. This work reveals that the cutaneous lesions of LSD provide the high-titer source required for acquisition of the virus by insects, thereby enabling the mechanical vector-borne transmission. IMPORTANCE Lumpy skin disease virus (LSDV) is a high consequence pathogen of cattle that is rapidly expanding its geographical boundaries into new regions such as Europe and Asia. This expansion is promoted by the mechanical transmission of the virus via hematogenous arthropods. This study quantifies the acquisition and retention of LSDV by four species of blood-feeding insects and reveals that the cutaneous lesions of LSD provide the high titer virus source necessary for virus acquisition by the insects. An artificial membrane feeding system containing a high titer of LSDV was shown to be comparable to a skin lesion on a clinically affected animal when used as a virus source. This promotes the use of these laboratory-based systems as replacements for some animal studies. Overall, this work advances our understanding of the mechanical vector-borne transmission of LSDV and provides evidence to support the design of more effective disease control programmes.

Larval morphology of the avian parasitic genus Passeromyia: playing hide and seek with a parastomal bar.

The enigmatic larvae of the Old World genus Passeromyia Rodhain & Villeneuve, 1915 (Diptera: Muscidae) inhabit the nests of birds as saprophages or as haematophagous agents of myiasis among nestlings. Using light microscopy, confocal laser scanning microscopy and scanning electron microscopy, we provide the first morphological descriptions of the first, second and third instar of P. longicornis (Macquart, 1851) (Diptera: Muscidae), the first and third instar of P. indecora (Walker, 1858) (Diptera: Muscidae), and we revise the larval morphology of P. heterochaeta (Villenueve, 1915) (Diptera: Muscidae) and P. steini Pont, 1970 (Diptera: Muscidae). We provide a key to the third instar of examined species (excluding P. steini and P. veitchi Bezzi, 1928 (Diptera: Muscidae)). Examination of the cephaloskeleton revealed paired rod-like sclerites, named 'rami', between the lateral arms of the intermediate sclerite in the second and third instar larva. We reveal parastomal bars fused apically with the intermediate sclerite, the absence of which has so far been considered as apomorphic for second and third instar muscid larvae. Examination of additional material suggests that modified parastomal bars are not exclusive features of Passeromyia but occur widespread in the Muscidae, and rami may occur widespread in the Cyclorrhapha.

Review of Synthesiomyia nudiseta (Diptera: Muscidae) as a useful tool in forensic entomology.

Synthesiomyia nudiseta (van der Wulp, 1883) is a synanthropic muscid found in tropical and subtropical zones around the world. The larvae of this species are a secondary agent of myiasis with necrophagous habits and play an important role in forensic entomology, as they are used as an indicator of post-mortem interval. Adults can be considered vectors of etiological agents such as Escherichia coli and Shigella dysenteriae. Due to its ability to adapt to different environmental conditions, its high dispersal capacity (shown by its introduction to Europe), its predatory habits in the last larval stage and the difficulty of identifying it, a very important goal is to update our knowledge about this species. Therefore, the main objective of this paper is to review the identification, geographical distribution and biology of this species in order to provide better support to investigations involving this fly.

Flies Exploit Predictable Perspectives and Backgrounds to Enhance Iridescent Signal Salience and Mating Success.

Communication requires both the encoding of information and its effective transmission, but little is known about display traits that primarily serve to enhance efficacy. Here we examined the visual courtships of Lispe cana, a cursorial fly that lives and mates in heterogeneous foreshores, and tested the prediction that males should seek to enhance signal salience and consequent fitness through the flexible choice of display locations. We show that courting males access the field of view of females by straddling them and holding their wings closed before moving ahead to present their structurally colored faces in ritualized dances. Males preferentially present these UV-white signals against darker backgrounds and the magnitude of contrast predicts female attention, which in turn predicts mating success. Our results demonstrate a striking interplay between the physical and attentional manipulation of receivers and reveal novel routes to the enhancement of signal efficacy in noisy environments.

A review and illustrated description of Musca crassirostris, one of the most neglected haematophagous livestock flies.

Tabanids, stomoxyine flies, hippoboscids and tsetse flies are the most well-known brachyceran biting flies of livestock. Only a few other higher Diptera have developed the unique mouthparts required for blood feeding. These neglected blood feeders can also have direct effects on hosts through blood loss, and are likely to contribute to the transmission of pathogens. Musca crassirostris (Diptera: Muscidae) is one of the most abundant of the muscid flies with this haematophagous lifestyle; it is widespread in the Palaearctic, Afrotropical and Oriental regions. The present study reviews and summarizes the biology and morphology of this species, and its potential for impact on animals and humans. The study also provides a fully illustrated description of the fly to facilitate its identification, and reviews information on abundance, with a focus on recent trapping surveys in Thailand. When sampled using traps designed for other biting flies, M. crassirostris appears to be four and 45 times more abundant than stomoxyines and tabanids, respectively. High numbers of M. crassirostris in the vicinity of livestock have also been associated with outbreaks of disease, such as that of a fatal plague in bovine farms in Egypt. This calls for a reconsideration of its potential impacts on livestock economics and health, and thus the development of suitable control methods.

Ethiopian muscids (Diptera, Muscidae) egg-carriers of Stylogaster Macquart (Diptera, Conopidae).

We here report for the first time on the presence of three species of the conopid genus Stylogaster Macquart (Diptera, Conopidae) in Ethiopia, viz. S. nitens Brunetti, S. westwoodi Smith and Stylogaster sp.. We further screened 908 muscid flies (Diptera, Muscidae) for the presence of impaled eggs of Stylogaster and recorded eggs on 89 individuals (9.8%). Eggs were impaled on eight species, viz. Limnophora translucida Stein, Musca lusoria Wiedemann, Musca splendens Pont, Neomyia chrysopyga (Emden), Pseudohelina nigritarsis (Jaennicke), Stomoxys omega Newstead, Stomoxys taeniatus Bigot and Stomoxys varipes (Bezzi). The maximum number of eggs found on a single muscid was six. We illustrated the dissected eggs. L. translucida, M. lusoria, M. splendens, N. chrysopyga and S. varipes are reported as new muscid hosts species for Stylogaster.

First record of Fanniidae (Diptera) from New Caledonia.

The family Fanniidae is newly recorded to New Caledonia. Adult male and female of Fannia capitalis Pont are illustrated as well as the peculiar male terminalia, the female ovipositor, and the egg. Male phallic process and egg are described for the first time.

Landmark and outline-based geometric morphometrics analysis of three Stomoxys flies (Diptera: Muscidae).

Adult flies of the genus Stomoxys Geoffroy, 1762 (Diptera: Muscidae), especially S. pullus Austen, 1909, S. uruma Shinonaga et Kano, 1966 and S. indicus Picard, 1908, are morphologically similar and sometimes difficult to distinguish when using external morphological characteristics. These species may act as vectors and/or potential vectors of many pathogens (virus, bacteria and protozoa). Their correct identification is important to target the vectors involved in the transmission of the pathogens and also helps in the fly control program.The aim of the present study was to distinguish three species which are difficult to separate using traditional diagnostic characters for species of Stomoxys such as colour patterns and body proportions. Modern morphometrics, both landmark and outline-based, was used to access wing geometry of S. pullus, S. uruma and S. indicus. A total of 198 and 190 wing pictures were analysed for landmark- and outline-based approaches, respectively. Wing shape was able to separate species and sexes of the three Stomoxys flies with highly significant difference of Mahalanobis distances. The cross-validated classification scores ranged from 76% to 100% for landmark and 77% to 96% for outline-based morphometrics. The geometry of wing features appears to be a very useful, low-cost tool to distinguish among the vectors S. pullus, S. uruma and S. indicus.

External Morphology of Stable Fly (Diptera: Muscidae) Larvae.

Scanning electron microscopy was used to examine the external morphology of first-, second-, and third-instar stable flies (Stomoxys calcitrans (L.)). In the cephalic region, the antennae, labial lobe, and maxillary palpi are morphologically similar among instars. Antennae comprise a prominent anterior dome that is the primary site of olfaction, while the maxillary palpi are innervated with mechano- and chemosensilla and scolopodia. The ventral organ and facial mask, also located in the pseudocephalon, are not well-developed in first instars, but become progressively more so in the subsequent instars. When the pseudocephalon is partially retracted, anterior spines cusp around the oral ridges of the facial mask. This indicates the anterior spinose band may be used in conjunction with the facial mask in predigestion. Functional anterior spiracles are absent on first instars, but become evident as a pair of palmate spiracular processes with five to seven lobes in second and third instars. A pair of Keilin's organs, functioning as hygroreceptors, is located on each thoracic segment. Abdominal segments are marked with ventral creeping welts, the anal pad, anus, papillae, and posterior spiracles. Ventral creeping welts are thought to aid in locomotion, while the anal pad acts as an osmoregulatory structure. Posterior spiracles are modified from round spiracular discs with two straight slits in the first instar to triangular discs with two and three sinuous slits in the second and third instars, respectively.

Morphology successfully separates third instar larvae of Muscina.

Three representatives of the muscid genus Muscina Robineau-Desvoidy (Diptera: Musicidae), Muscina levida (Harris), Muscina prolapsa (Harris) and Muscina stabulans (Fallén), are well known for their medical, veterinary and forensic importance. However, data in the literature provide contradictory information for the identification of third instar larvae of these species. This hinders easy species differentiation because it requires the rearing of material to adult stages in order to facilitate reliable identification. Third instar larvae of these three Muscina species were studied in detail using light and scanning electron microscopy to provide thorough redescriptions of their morphology. Existing information concerning third instar morphology, with particular reference to its value for taxonomy and identification, is revised and discussed. Emden's spiracular distance factor (SDF) is considered here as inappropriate for identification purposes in third instar larvae of Muscina spp. because its values are not constant ratios but increase during larval maturation and overlap in the examined species. These species were, instead, discriminated here by differences in the spinulation patterns of their abdominal segments.

Descriptions of six new species of Phaonia Robineau-Desvoidy (Diptera: Muscidae) from China.

This paper provides diagnoses and keys to species in the Phaonia fuscicoxa-group and the Phaonia barkama-group from China; describes six new species, namely Phaonia subfuscicoxa Xue and Rong, sp nov: , Phaonia hypotuberosurstyla Xue and Rong, sp nov: , Phaonia caesiipollinosa Xue and Rong, sp nov: , Phaonia daliensis Xue and Du, sp nov: , Phaonia quadratilamella Xue, sp nov: , and Phaonia maoershanensis Xue, sp nov: We report the distributions and provide notes on the affinities of known species.

Keys to the species of Mydaeinae (Diptera: Muscidae) from China, with the description of four new species.

Four new species of Mydaeinae, Mydaea franzosternita n sp., Myospila apicaliciliola n sp., Myospila maoershanensis n sp., and Myospila subflavipennis n sp., are described and illustrated here for the first time. A key to the genus of Mydaeinae from China and keys to species of genera from Mydaeinae are provided.

Contributions to the taxonomy and faunistics of some arctic species of Spilogona Schnabl (Diptera: Muscidae).

New synonymies in the genus Spilogona Schnabl, 1911 are proposed: S. novaesibiriae (Frey, 1915) = S. obsoleta (Malloch, 1920), syn. nov.; S. trianguligera (Zetterstedt, 1838) = S. setinervis (Huckett, 1932), syn. nov. Spilogona trianguligera and S. tendipes (Malloch, 1920) are newly recorded from Greenland, and S. trianguligera also from W Siberia. The female of S. vikhrevi Sorokina, 2010 is described and the species in newly recorded from Wrangel Island (Far East Russia).

An illustrated identification key to early instar larvae of forensically important Muscidae (Diptera) of the western Palaearctic region.

There is a significant gap in the availability of comprehensive identification keys for the early larval stages of forensically important fly species. While well-documented identification keys exist for the third instar larvae, particularly for the Calliphoridae, Muscidae and Sarcophagidae families, there is a notable scarcity of keys for the first, except Calliphoridae, and the second instar larvae, with no such resources available for muscid species. The second instar larvae suffer the most from the lack of morphological descriptions and available identification keys. The Muscidae is one of the most frequently reported dipteran families of forensic importance colonising animal cadavers and human corpses. Nevertheless, descriptions of the morphology of their early instars remain scarce and limited to only a few species, thus their larval identification is challenging or impossible. Considering the numerous challenges associated with studying small-sized entomological material, we tested whether it is feasible to identify muscid flies to the species or at least genus level based predominantly on the details of the cephaloskeleton. To overcome the obstacle of observing details of small sclerites, especially their shapes and interconnections, we effectively employed confocal laser scanning microscopy (CLSM) as a supplementary method for light microscopy (LM). This study provides an identification key for first and second instar larvae of forensically important muscid species from the western Palaearctic (Europe, North Africa, Middle East). The proposed key primarily utilises details of the cephaloskeleton with only addition of external morphology.

Molecular characterization and immunolocalization of the olfactory co-receptor Orco from two blood-feeding muscid flies, the stable fly (Stomoxys calcitrans, L.) and the horn fly (Haematobia irritans irritans, L.).

Biting flies are economically important blood-feeding pests of medical and veterinary significance. Chemosensory-based biting fly behaviours, such as host/nutrient source localization and ovipositional site selection, are intriguing targets for the development of supplemental control strategies. In an effort to expand our understanding of biting fly chemosensory pathways, transcripts encoding the highly conserved insect odorant co-receptor (Orco) were isolated from two representative biting fly species, the stable fly (Scal\Orco) and the horn fly (Hirr\Orco). Orco forms a complex with an odour-specific odorant receptor to form an odour-gated ion channel. The biting fly transcripts were predicted to encode proteins with 87-94% amino acid similarity to published insect Orco sequences and were detected in various immature stages as well as in adult structures associated with olfaction, i.e. the antennae and maxillary palps, and gustation, i.e. the proboscis. Further, the relevant proteins were immunolocalized to specific antennal sensilla using anti-serum raised against a peptide sequence conserved between the two fly species. Results from the present study provide a basis for functional evaluation of repellent/attractant effects on as yet uncharacterized stable fly and horn fly conventional odorant receptors.

Larval morphology, development and forensic importance of Synthesiomyia nudiseta (Diptera: Muscidae) in Europe: a rare species or just overlooked?

The muscid Synthesiomyia nudiseta (van der Wulp, 1883) is a species with forensic importance in tropical and subtropical regions of the world. This fly has recently been introduced into southern Europe and, until now, had not been recorded in forensic cases in this region. Here, morphology of all larval instars of S. nudiseta is documented in detail by using a combination of light and scanning electron microscopy. Literature data concerning larval morphology are revised and characters allowing identification from other forensically important Muscidae are listed. The life cycle of this species was studied at four constant temperatures: 15, 20, 25 and 30 °C. Total development varied between 46.50 ± 0.97 days at 15 °C and 15.39 ± 0.32 days at 30 °C. Moreover, we report this species breeding in human corpses, for the first time in Europe, in forensic cases from autopsies at the Anatomical Forensic Institute of Madrid and the Institute of Legal Medicine of Alicante, Spain.

A study of the Phaonia angelicae group (Diptera: Muscidae), with descriptions of six new species from China.

The P. angelicae group from China was studied, and six new species are described: P. hanmiensis, n. sp., P. nanlingensis, n. sp., P. reduncicauda, n. sp., P. spargocerca, n. sp., P. subincana, n. sp., and P. varimargina, n. sp. A key to the identification of males of the 40 Chinese species is given.

Sequence analysis of the internal transcribed spacer 2 (ITS2) from Philornis seguyi (García, 1952) and Philornis torquans (Nielsen, 1913) (Diptera: Muscidae).

Philornis Meinert, 1890 (Diptera: Muscidae) is a genus of Neotropical dipterans that parasitise birds. The currently used external morphological characters to distinguish between species within this genus present some limitations. We used the second internal transcribed spacer region (ITS2) of the rRNA gene as a molecular marker to differentiate adult specimens of Philornis identified morphologically as Philornis torquans and Philornis seguyi from different localities. Specimens identified as P. seguyi from Magdalena (Buenos Aires Province) showed an ITS2 sequence different from that for P. torquans, whereas all other specimens of P. seguyi had sequences identical to those for P. torquans. These findings do not necessarily confirm that specimens from Magdalena indeed belong to P. seguyi, nor that P. seguyi is a valid species. Instead, they alert us about the potential for species misidentification when using morphological characters alone. The use of molecular approaches to aid the identification of Philornis spp. will shed light on the systematics of this group. P. torquans is reported for the first time in Mendoza Province and Uruguay.

New species and redescriptions of the New Zealand genus Exsul Hutton (Diptera: Muscidae: Coenosiinae).

Exsul Hutton (Diptera, Muscidae) is revised and the genus diagnosis is enlarged. The female of Exsul singularis Hutton is described for the first time. The male and female terminalia of all species are described and illustrated. Exsul alfredoi sp. n. is described and illustrated and compared with the other known species. A key to separate the species is given.

The identity of the genus Scatocoenosia Schnabl, 1915 (Diptera: Muscidae).

A re-description is given of the problematic genus and species Scatocoenosia cordyluraeformis Schnabl, 1915. Illustrations of the male and female terminalia, and images of the female holotype and the male of this species, are given. The syno-nymy of Scatocoenosia Schnabl, 1915 with Spilogona Schnabl, 1911 is confirmed, and notes are given on the relationships of S. cordyluraeformis with other Spilogona species.