Comparative scanning electron microscopy of third-instar Hypoderma spp. (Diptera: Oestridae)

Scanning electron microscope study of third-instar larvae of four species of Hypoderma revealed differences among species in the pattern of spination, spine morphology and morphology of the spiracular plates. These observations identify characters that enable the differentiation of Hypoderma actaeon and H. diana, parasitizing red deer ( Cervus elaphus ) in Europe, and provide additional characters for differentiating H. bovis and H. lineatum parasitizing cattle.     

Cuticular sensilla on newly hatched larvae of Cuterebra fontinella Clark (Diptera : Cuterebridae) and Hypoderma spp. (Diptera : Oestridae)

Cuticular sensilla on newly hatched larvae of Cuterebra fontinella Clark (Diptera : Cuterebridae), Hypoderma lineatum (de Vill.) and H. bovis (L.) (Diptera : Oestridae) were studied by scanning electron microscopy. Three basic types of sensilla were found on C. fontinella larvae, while only 2 were present on the larvae of Hypoderma spp. Sensilla were more numerous on the C. fontinella larvae than on the Hypoderma spp. larvae (210 vs 66). There were no noticeable differences between the sensory complements of H. lineatum and H. bovis. Comparison of the number and type of sensilla on the parasitic larvae with those on free-living muscomorph Diptera larvae indicate that increasing numbers and greater morphological diversity correlate with increasing requirements for location of a suitable environment in which to feed and develop.     

Scanning electron microscopy of Oestrus ovis larvae (Diptera: Oestridae): skin armour and posterior spiracles

The skin armour and posterior spiracles of L1, L2 and L3 of Oestrus ovis (Diptera: Oestridae) are described and documented photographically by a scanning electron microscope. The complex and variable morphology of the attack organs of larvae of different stages makes it easier to understand better their exceeding adaptability to the host. The SEM survey has also allowed an apparently unreported anatomic particular with a probable sensory function to be detected in L1.     

Cuticular structures on the antennae of Hypoderma bovis De Geer (Diptera : Oestridae) females

The antennae of Hypoderma bovis (Diptera: Oestridae) females were examined using scanning electron microscopy. Each antenna is composed of 3 parts: the scape, the pedicel, and the funiculus, with a large, protruding arista. Mechanoreceptors are found on the proximal and lateral margins of the scape and pedicel, respectively. Microtrichia, which are presumably non-innervated, are located evenly over both the outer surface of the scape and the inner surface of the pedicel. A narrow band of microtrichia is present proximally on the outer surface of the pedicel. The entire funicular surface is densely covered with microtrichia. Small patches lacking these microtrichia appear as depressions or “pits” (8–20 μm in diameter) on the surface of the funiculus. Olfactory sensilla found on the funicular surface include basiconica type 1, basiconica type 2, and trichoid sensilla. The sensilla basiconica commonly occur in pits on the anterodorsal surface of the funiculus. Trichoid sensilla are abundant on the posteroventral surface of the funiculus and do not appear to occur in pits. In addition, clavate and peg sensilla, whose functions are unknown, are found in low numbers on the funicular surface. There may be as many as 300 olfactory pits on the anterodorsal surface of each funiculus. These are single-chambered and contain 6 or fewer sensilla basiconica. We propose that a relatively high number of pits may be characteristic of flies in Superfamily Oestroidea (as compared with those of Superfamily Muscoidea), but that pit morphology within the Calyptratae is not Superfamily-specific.     

在藏羚羊上发现的中国第四种皮蝇

对于从可可西里藏羚羊上采集到的藏羚羊皮蝇Hypoderma sp.3龄幼虫进行了形态学观察,发现其在伪头、第10腹节上的刺、气门板形态上与中国现有记录的3种皮蝇（牛皮蝇、纹皮蝇和中华皮蝇）有着明显的区别。对藏羚羊皮蝇的线粒体COI 基因种属特异性序列研究表明,其与牛皮蝇、纹皮蝇和中华皮蝇对应序列的相似性分别为88.1％,87.8％和88.8％。据此认为采自藏羚羊的此种皮蝇可能为中国境内又一皮蝇新种或新记录种,为中国第四种皮蝇。     

Mating behavior and thermoregulation of the reindeer warble fly,Hypoderma tarandi L. (Diptera: Oestridae)

Hypoderma (=Oedemagena) tarandi L. (Diptera: Oestridae) is characterized by a mating strategy in which both sexes meet and mate at two types of distinct topographical landmarks. In the expansive, treeless vidda (= tundra-like) biome, mating places are unique, rocky areas located along rivers and streams or in rocky areas of drying river and stream beds. In wooded valleys below the vidda, flies mated at certain topographical areas along dirt road tracks/paths. Thermoregulatory activities of males occupying perches at mating places included selection of substratum at perch site, orientation of body to sun's rays, crouching, stilting, and flights into upper cooler air. On warm sunny days males perched for just 1–2 min before flying up into cooler air to promote cooling. Laboratory and field studies revealed that flies could not metabolically cool down when held at 25–38°C. Time spent at mating places depended on temperature, duration of sunshine, and wind velocity. Males were very aggressive in pursuing allHypoderma-sized objects that passed by them or that landed near them, but they did not defend specific perch sites. Males either pursued and caught females in flight, or they hopped onto females that landed near them. During 5 years, 74 males and 14 females were seen at mating places. Dissection of six females caught at mating places revealed them to be recently eclosed flies full of fat body and with all eggs intact; two not paired with males were non-inseminated. Three experimentally paired females remainedin copulo for 10, 13, and 19.5 min.     

Early detection of cattle grub (Hypoderma lineatum and H.bovis) (Diptera, Oestridae) using ELISA

An enzyme-linked immunosorbent assay (ELISA) to detect cattle grub-infested animals in the autumn in Alberta has been developed. Antibody to Hypoderma lineatum de Vill. was detectable as early as 6 weeks post-infestation in artificially infested steers. Peak antibody concentrations preceded the peak in maximum 'apparent' grub numbers which occurred between 37 and 43 weeks after infestation. Natural infestations with H. lineatum and H. bovis L., ranging from one to ninety-two grubs per calf, were readily detected by November and the incidence of false positives in uninfested calves was only 5%. Low level (1-4 grubs) infestations of H. bovis only became detectable in February with peak antibody concentrations occurring at the end of April. Prevalence of cattle grub infestations in southern Alberta was shown to be 37% during this study.     

Identification of Lutzomyia spp. (Diptera: Psychodidae) verrucarum group through electron microscopy of its eggs

The value of Colombian phlebotomine eggs for species determination was studied with a scanning electron microscope. The species diversity and medical importance of the verrucarum group were the bases to select Lutzomyia youngi, Lutzomyia evansi, Lutzomyia columbiana and Lutzomyia longiflocosa. The egg surface was poligonal. Lutzomyia youngi, and Lutzomyia columbiana had pentagonal or hexagonal patterns; Lutzomyia evansi elongated polygons and Lutzomyia longiflocosa irregular polygonal sculpturing, frequently rectangular. Egg scanning electron microscopy is reliable to identify species of the verrucarum group.     

Phylogeny of Oestridae (Insecta: Diptera)

The phylogeny of Oestridae was analysed at the generic level using 118 characters from all developmental stages and including morphology, ontogeny, physiology and behaviour. Four major clades were given subfamilial rank with the phylogenetic relationship (Cuterebrinae (Gasterophilinae (Hypodermatinae + Oestrinae))). The subdermal parasites of the African elephant, Neocuterebra squamosa Grünberg and Ruttenia loxodontis Rodhain, had their most probable affiliation subordinate to the clade of stomach parasites, although their exact position needs further investigation. Genus Ochotonia Grunin, which is known from a single third‐instar larva only, was the probable sister group of all other Hypodermatinae. Twenty‐five oestrid genera were recognised as valid and those containing more than one species were defined through lists of autapomorphies. Cuterebra Clark was proposed as a senior synonym of Alouattamyia Townsend, Andinocuterebra Guimarães, Pseudogametes Bischof and Rogenhofera Brauer. The clade of hypodermatine ungulate parasites (Hypoderma Latreille +Pallasiomyia Rubtzov +Pavlovskiata Grunin +Przhevalskiana Grunin +Strobiloestrus Brauer) remained largely unresolved, and genus Przhevalskiana emerged without defining characters.     

Morphology of the second- and third-instar larvae of Dermatobia hominis by scanning electron microscopy

Larvae of Dermatobia hominis 10–27 days old were collected from experimentally infected rats and their morphology was studied by scanning electron microscopy. The moult from the second to third instar occurs at 18 days, with emergence from the host at 30 days post-infection. The second-instar larvae bear on the pseudocephalon, antennae (coeloconic sensilla), and coeloconic and basicoconic sensilla on the maxillary sensory complex. The thoracic segments bear small backwardly-directed spines anteriorly and ventral trichoid and campaniform sensilla. The first four abdominal segments have small and large backwardly-directed spines that are absent on segments five and six. The seventh and eighth abdominal segments have medium-sized forwardly-directed spines. Abdominal segments are encircled by campaniform sensilla. The terminal end of the eighth abdominal segment bears the anus, prominent anal lobes and two spiracular openings on each spiracular plate. Spiracular plates show a radial sun ray pattern. The rear abdomen also bears an ecdysal aperture, several pores and eight coeloconic sensilla. Although there are slight morphological differences, the spines (predominantly flat and thorn-like) and sensilla (campaniform and coeloconic) of the third-instar larvae show a similar arrangement to that of second-instar larvae. Thoracic trichoid sensilla are not seen in third-instar larvae. A perispiracular gland aperture is situated above each posterior spiracular opening. These morphological features are compared with those of other cuterebrid larvae.     

Effects of Cephenemyia spp. (Diptera: Oestridae) on the nasopharynx of black-tailed deer (Odocoileus hemionus columbianus)

A study was conducted to determine gross and microscopic tissue changes in the nasopharynx of black-tailed deer (Odocoileus hemionus columbianus) infected with nasal bot fly larvae (Cephenemyia spp.). Paired retropharyngeal recesses were the preferred sites for the growing second and third stage larvae of two species of Cephenemyia (C. apicata and C. jellisoni). Retropharyngeal recesses distended into "pouches" that harbored up to 30 larvae. Pouches were oriented caudal-laterally toward the basisphenoid bone of the cranium. Lateral support of the pouch mass was provided by the stylohyoid bone. The laryngeal orifice was never occluded by the enlarged recesses. The distal pouch wall was relatively thin and remained uniform in thickness as expansion progressed. Occasionally, aberrant larvae were found protruding through the distal wall of the pouch. Disruption of the epithelium and submucosa by larval mouth hooks and integumentary spines were examined by scanning electron microscopy. Histological examination of infected recesses revealed substantial loss of epithelium and mucous glands. Enlargement of recesses into pouches resulted from fibrosis. Healing occurred after larvae egressed from the pouches. Degenerating mucous glands, epithelial metaplasia, epithelial desquamation, and intense inflammation were found near larvae. An eosinophilic exudate with a mixture of macrophages and erythrocytes was present in the lumen of the pouch. The presence of larvae within the pouch inhibited secondary bacterial infection and suppuration. Infection by larvae caused severe local trauma and intense tissue response.     

A third species of Hypoderma (Diptera: Oestridae) affecting cattle and yaks in China: molecular and morphological evidence

Cattle and yak hypodermosis in China is caused by Hypoderma bovis and H. lineatum, with a prevalence reaching up to 98-100% of the animals and maximum intensities exceeding 400 warbles for each animal. A third species, H. sinense, is also considered by Chinese researchers to affect livestock. The molecular characterization of the most variable region of the mitochondrial cytochrome oxidase I gene and of the ribosomal 28S gene has been performed for the third-stage larvae collected from cattle and yaks in China and identified (on the basis of the spinulation on the ventral side of the 10th segment) as H. bovis, H. lineatum, and H. sinense. Amplicons were digested with the HinfI and BfaI restriction enzymes, which provided diagnostic profiles to simultaneously differentiate the 3 Hypoderma species. Third-stage larvae of H. sinense were also examined by scanning electron microscopy, which revealed proper morphological characteristics different from those of H. bovis and H. lineatum. The molecular and morphological evidence herein reported support the existence of a third species of Hypoderma affecting cattle and yaks in China, and the results provide new tools for unequivocal identification of this species and present key components for the evaluation of its endogenous cycle and pathogenicity in animals and humans.     

In vitro rearing Oestrus caucasicus third-instar larvae and pupae (Diptera: Oestridae) from naturally-infested Iberian ibex, Capra Pyrenaica (Artiodactyla: Bovidae)

Third-instar Oestrus caucasicus larvae (n = 236) obtained from Iberian ibex, Copra pyrenaica, were reared in a laboratory to obtain adult flies. They were maintained at a temperature of 21.9 +/- 2.7 degrees C and a relative humidity of 38.9 +/- 8.0 %. In all, 78 imagos emerged (33.1 %), with a sex-ratio at emergence not differing significantly from 1:1; 25 larvae did not complete pupariation. A total of 14 adult flies (17.9 % of the adults obtained) showed malformations, mainly in their wings. The pupariation period lasted around 30 hours and the pupal stage lasted on average 29.8 +/- 6.8 days. The success of pupation in both sexes was mainly determined by the weight of the larvae. Sexual dimorphism, with higher weights in females, was evident in third-instar arvae, pupae and adults. The mean longevity of adult flies was 224.8 +/- 91.4 hours and males generally survived for onger than the females.