Female Black Flies Research Articles

Dipteran predators of Simuliid blackflies: a worldwide review.

Haematophagous female blackflies (Diptera: Simuliidae) are serious biting pests and obligate vectors of vertebrate pathogens, namely filarial Dirofilaria, Mansonella, Onchocerca and protozoal Leucocytozoon. Immature stages of Simuliidae inhabit lotic waterways, the sessile larvae filter-feeding and often forming a large proportion of the benthic biomass, usually aggregated in well-oxygenated sections of streams, rivers, waterfalls and spillways. Simuliid control practices depend on larvicidal chemicals, biological products (bacteria, nematodes) and environmental modification. The potential use of predators for biological control of Simuliidae has not been exploited. Predators of Simuliidae include examples of at least 12 families of Diptera and other predaceous arthropods (Crustacea and insects: Coleoptera, Odonata, Plecoptera, Trichoptera), invertebrates (notably Turbellaria), as well as browsing fish. Diptera impacting upon simuliid populations comprise mainly Chironomidae, Empididae and Muscidae, although several other families (Asilidae, Dolichopodidae, Phoridae, Drosophilidae, Scathophagidae) play a significant role as predators. Details of predator and prey species and life stages are presented, by zoogeographical region, including the prevalence of cannibalism among Simuliidae.

NATURAL INFECTIONS WITH FILARIAL LARVAE IN TWO SPECIES OF BLACK FLIES(DIPTERA: SIMULIIDAE) IN NORTHERN THAILAND.

To find out the natural infection with filarial larvae, female adult black flies were collected on a human attractant in December, 2001 at Tambol Ban Laung (altitude 750 m), Doi Inthanon National Park, in northern Thailand. The total number of females collected was 823 : of which 557 (67.7%) were identified as Simulium asakoae Takaoka et Davies, 144 (17.5%) as S. nigrogilvum Summers, 97 (11.8%) as S. nakhonense Takaoka et Suzuki and 25 (3%) as other six simuliid species. By dissections, eight third-and one second-stage larvae of unidentified filarial species were found in one of 138 S. nigrogilvum and one of 484 S. asakoae, respectively. Non-filarial nematodes were found in 1.03% (5/484) of S. asakoae. This is the first report of natural infections of two black-fly species, S. nigrogilvum and S. asakoae, with a filarial larva.

Immunolocalization of vesicular stomatitis virus in black flies (Simulium vittatum).

Vesicular stomatitis, a disease of cattle, horses, and swine, is caused by either vesicular stomatitis virus, New Jersey serotype (VSV-NJ), or vesicular stomatitis virus, Indiana serotype, which are related viruses in the genus Vesiculovirus, family Rhabdoviridae. Although recognized for at least 160 years, the epidemiology and pathogenesis of this disease remains undefined. Black flies have been suggested as a vector for VSV-NJ. In this study we infected three- to four-week-old female black flies with VSV-NJ via feeding of virus-spiked ox blood or intrathoracic inoculation, and demonstrated the location of virus by immunohistochemistry. These preliminary findings suggest that VSV-NJ initially infects the gut in the natural situation but that subsequent spread to the salivary gland may be blocked in older flies, decreasing their ability to transmit the virus. The pattern of staining was different in intrathoracic inoculated flies. In these flies, salivary gland involvement was more likely, and extensive staining of eye, brain, and hemolymph suggested a more generalized infection that apparently circumvented the gut. We conclude that intrathoracic inoculation may be an inappropriate method of infection for determining vector competence and that the age of the vector should be considered when conducting competency studies.

Humoral immune response of Simulium damnosum s.l. following filarial and bacterial infections.

The time-course of the humoral immune response of female blackflies after a challenge with bacteria, different Onchocerca microfilariae species, bacterial endotoxin and microfilarial extract was investigated. Strong bacteriolytic and growth inhibition activities against the Gram-positive bacterium Micrococcus luteus were induced by all agents. Specific differences were found in activity levels and time-course. Notably the endotoxin lipopolysaccharide (LPS) induced a very early, profound bacteriolytic and antibacterial response, which declined within a day after injection. In contrast, the bacteriolytic activities after Escherichia coli D31 and Onchocerca microfilariae infections were lower, but remained elevated over the observation period of 4 days. The bacteriolytic activity was correlated to a haemolymph protein with a molecular weight of around 14 kDa. Anti-Gram-positive activity in the E. coli infected group appeared within the first 6 h. However, it took 4 days in the microfilarial infected blackflies to reach significant levels. The active agent was identified to be a peptide with a molecular weight of around 4-4.5 kDa. Activity against the Gram-negative bacteria E. coli was detected in blackflies injected with E. coli D31, O. dukei microfilariae and microfilarial extract on days 1 and 4 after injection. The immune response in S. damnosum s.l. naturally infected via a bloodmeal on cattle supported the findings of the experimental infections. Similarities of the immune response kinetics between bacterial and filarial infections suggested that intracellular Wolbachia bacteria, released from microfilariae, could be responsible for the antibacterial response. This is supported by the observation that the induction of an immune response in the Drosophila melanogaster mbn-2 cell line by the filarial extract is blocked by polymyxin B, which forms inactive complexes with bacterial LPS.

Suppression of mosquito (Diptera: Culicidae) and black fly (Diptera: Simuliidae) blood feeding from Hereford cattle and ponies treated with permethrin.

The blood feeding of mosquitoes and black flies from Hereford cattle and ponies treated with commercial formulations of permethrin was evaluated using an animal enclosure trap sample system that allowed comparison of insect blood-feeding levels between treated and nontreated animals. Blood feeding of both Aedes dorsalis Meigen and A. melanimon Dyar from heifers treated with pour-on concentrate and whole body spray treatments was reduced significantly by 79-88% at 4 d posttreatment, with apparent but not significant reductions of 61-68% at 11 d posttreatment. Simulium bivittatum Malloch and S. griseum Coquillett blood feeding was reduced significantly by 96% to >99% at 4 d posttreatment, but apparent reductions of 30-87% at 11 d posttreatment were not significant. Blood feeding of S. bivittatum from ponies treated with a permethrin fly wipe was reduced significantly by 98 and 87% at 1 and 7 d posttreatment, respectively. No evidence of treatment-induced mortality was observed for recently blood-fed female mosquitoes or black flies captured from treated animals and held for 24 h. The potential benefit of using permethrin to protect livestock from insect-transmitted pathogens was estimated with a model based on level of host attack, pathogen infection rate in the vector, and suppression of blood feeding. Suppression of blood feeding by 90% is predicted to prevent the exposure of a host to a pathogen for up tolO d at 1,000 insect feedings per d when the vector population infection rate is one insect per 1,000. If insect feedings are lower (100/d) and the insect infection rate remains at one per 1,000, protection is predicted for 100 d. In contrast, a 90% suppression of blood feeding is predicted to provide protection for less than 1 d at 1,000 feeding per day and a vector infection rate of one insect per 100.

Biting behaviour and potential vector status of anthropophilic blackflies in a new focus of human onchocerciasis at Minaçu, central Brazil.

Monthly collections were made of man-biting female blackflies: Simulium auripellitum Enderlein, S. guianense Wise, S. minusculum Lutz and S. nigrimanum Macquart (Diptera: Simuliidae) from four catching stations in the newly discovered focus of human onchocerciasis at Minaçu (13 degrees 35 minutes S 48 degrees 18 minutes W), 300 km north of Brasília in Goiás State. These provided baseline data on biting habits, population density and seasonal prevalence during the year before completion of the Serra da Mesa hydroelectric dam on the Rio Tocantins near Minaçu, in a project investigating the effect of dam construction on onchocerciasis transmission in the area. All four simuliid species were most abundant during the dry season, and only bit in low numbers (S. auripellitum S. minusculum, S. nigrimanum) or were absent (S. guianense) in the wet season. Simulium minusculum was the predominant species at all catching stations, being particularly abundant by the large River Tocantins. The other three species were mainly associated with smaller rivers. In the dry season, biting rhythms of S. minusculum varied with catching site, while S. nigrimanum showed peaks of activity in early morning and during the afternoon. Experimental infection with Onchocerca volvulus (Leuckart) (Nematoda: Onchocercidae), from a human volunteer, showed that this parasite could develop fully in the four simuliid species, which are all considered to be potential vectors in the area.

Do gravid black flies (Diptera: Simuliidae) oviposit at their natal site?

We present information on an aniline dye marking method for black flies. In the laboratory, adults were sprayed with 2% aqueous solutions of four colors of aniline dyes; brilliant blue and methyl orange gave the best results in longevity trials. In field trials we were able to recapture 1.3% of newly emerged marked and released flies at oviposition sites. Mark–release–recapture experiments were designed to distinguish among three competing models concerning oviposition site selection by gravid female black flies: (1) larval site fidelity (“Do flies return to the site that they experienced as larvae?”), (2) adult site fidelity (“Do flies return to the site that they experienced as adults?”), and (3) no site fidelity (“Do flies oviposit at random, i.e., without regard to adult or larval experience?”). Models 1 and 2 were rejected. There is, however, no reason to reject Model 3, the no site fidelity model. Thus, we conclude that for members of the S. venustum/verecundum complex (i.e., S. rostratum, S. venustum, and S. truncatum) females find an “apparently suitable” waterway in which to oviposit; this may or may not be their natal site.

Regulation of mating behavior by nutrition and the corpus allatum in both male and female Phormia regina (Meigen)

Both dietary protein and the corpus allatum (CA) were required for normal mating behavior in both male and female black blow fly, Phormia regina (Meigen). Nutrition (protein diet) activated the CA in both sexes. More than 10 mg of dietary liver was required for each male to result in 80% insemination of females, while 20 mg of liver was required for each female to allow 78% of females to become inseminated. Between 10 to 15 mg of protein meal (i.e., liver) was required to activate sexual receptivity in 71% of the females, while between 15 to 20 mg of liver was needed to support full oöcyte development in 70% of the females (Yin, C.-M., Zou, B.-X., Li, M.-F., Stoffolano, J.G., Jr. 1994. Discovery of a midgut peptide hormone which activates the endocrine cascade leading to oogenesis in Phormia regina Meigen. Journal of Insect Physiology, 40, 283–292). Allatectomy suppressed mating behavior more than 2-fold in both sexes. Topical application of 10 μg of S-methoprene (a juvenile hormone analogue) at 12 h after the onset of liver feeding restored sexual activity of both allatectomized males and females. Incidence of successful insemination increased as the oöcyte development progressed. Ovariectomy suppressed sexual receptivity more than 3-fold in liver-fed females. Thus, in addition to nutrition and the CA, ovaries and their developmentaal status can also affect the sexual receptivity in female P. regina.

Bite transmission of vesicular stomatitis virus (New Jersey serotype) to laboratory mice by Simulium vittatum (Diptera: Simuliidae).

Laboratory-reared female black flies (Simulium vittatum Zetterstedt) were infected experimentally with a 1997 vesicular stomatitis virus New Jersey serotype isolate and allowed to feed on susceptible laboratory mice. All mice exposed to black fly bite seroconverted by day 21 after infection, an indication of virus transmission. In addition, viral RNA was detected in the spleen of several mice. These findings are consistent with the hypothesis that black flies are involved in VSV-NJ transmission during epizootics in the western USA and represent the 1st confirmed example of biological transmission of an arbovirus by a member of the Simuliidae using an animal model.

Nectar versus honeydew as sources of sugar for male and female black flies (Diptera: Simuliidae).

Black flies were collected twice daily using a vehicle-mounted insect net on 8 sample days between 20 May and 9 July 1993. The contents of the crops and midguts of 160 male and 160 female Simulium venustum Say were analyzed individually by thin-layer chromatography. Fourteen different combinations were identified of the following sugars: fructose glucose sucrose/turanose, melezitose, raffinose, and stachyose. Presence of melezitose and stachyose indicated that black flies had fed on homopteran honeydew. Significantly more female (40.0%) than male flies (27.5%) fed on honeydew, and this difference was caused by a skewedness in the morning samples.

Sugar-meal sources used by female black flies (Diptera: Simuliidae): a four-habitat study

Adult black flies were sampled by sweep-netting vegetation in four habitats within Algonquin Provincial Park, Ontario: Davies Bog, the airfield, deciduous habitat, and coniferous habitat. Sugars in the crops and midguts of female flies (n = 773) were tested by thin-layer chromatography to determine whether the flies had fed on nectar or homopteran honeydew. Melezitose and stachyose were used as honeydew-indicator sugars. For Simulium venustum, it was found that significantly fewer black flies (19%) from the airfield contained honeydew sugars than black flies from the other three sites (34% from Davies Bog; 36% from deciduous habitat; 25% from coniferous habitat). We argue that black flies will feed on nectar or honeydew according to availability.

Control of pupal fat body disappearance in the female black blow fly, Phormia regina (Meigen) by the brain and the corpus allatum

Neck-ligation, brain implantation, allatectomy, methoprene treatment, and ovariectomy indicated that the disappearance of pupal fat body cells in newly emerged adult female blow fly, Phormia regina, is controlled by the brain and the corpus allatum (CA). Absence of brain or CA greatly lowered the rate of fat body cell disappearance (i.e. death). Dependency on the CA decreased from 0 to 36 h post-emregence, indicating that the CA was active during the earlier part of this timespan. Methoprene treatment enhanced pupal fat body cell disappearance in allatectomized females. Brain implantation restored the rate of pupal fat body cell disappearance in neck-ligated flies. Brains from day 1 sugar-fed flies proved to be more effective than those from day 2 sugar-fed flies, indicating that there may be a window after adult emergence that allows the brain to act directly or indirectly on the death of pupal fat body cells. Ovariectomy did not alter the rate of pupal fat body cell death in test animals. Dying pupal fat body cells were smaller in size, less dense (i.e. did not sink in saline like normal pupal fat body cells), and stickier (i.e. attached to other tissues tighter) than the healthy cells. A possible role played by ecdysteroids is also discussed.

Onchocerciasis in an expatriate living in Cameroon.

Travelers to West, central and eastern Africa as well as to selected areas of Latin America are at risk for infection with Onchocerca volvulus. Infection with this tissue nematode may cause chorioretinitis and keratitis, and it is responsible in endemic areas for blindness in millions.1 In addition to ocular manifestations, it can produce a distressing pruriginous dermatitis or subcutaneous nodules. Clinical manifestations vary according to the parasitic load, previous immunity, and duration of infection.1 Infection is initiated by inoculation with larvae during the bite of the Simulium black fly. Once in the connective tissue, larvae mature to filiform adults and may remain in tissues for years, with the clinical manifestations being produced by the inflammatory reaction to dying parasites. Female adults produce large amounts of microfilaria that migrate through skin and connective tissue; once an infected host is bitten, the infectious larvae develop again in the female Simulium black fly and the life cycle is completed. Transmission is from person to person, and may occur even after a relatively short exposure.2 Although infection of travelers with O. volvulus is rare, according to the Centers for Disease Control and Prevention, 46 new cases of onchocerciasis were diagnosed in the United States in 1994 (Dr. David Addis, personal communication, June 1996). We describe the case of an expatriate who became infected with O. volvulus and we review the treatment and recommendations for prevention of this parasitic infection.

Human individuals vary in attractiveness for host-seeking black flies (Diptera: Simuliidae) based on exhaled carbon dioxide.

The attractiveness of 5 male subjects was compared to determine if female black flies were attracted more readily by certain individuals and, if so, to determine the bases of such differences. During 2 yr, in experiments contrasting different groupings of the subjects, 1 subject was consistently the most attractive, whereas another was consistently the least attractive. Attractiveness of the remaining 3 subjects was intermediate. Removing exhaled breath resulted in a decrease of attractiveness to approximately 15% of previous levels and removed differences among subjects. Replacing breath exhalations with subject-specific CO2 levels restored subject attractiveness to approximately 90% of original levels. Exhaled breath from which CO2 had been removed with a soda-lime filter and then augmented with bottled CO2 was consistently more attractive than CO2 alone at the augmentation rate, indicating the role of other breath components. A human form plus breath odor was more attractive than a human form and a human form plus body odor. Results lend credibility to popular notions that some people are more attractive for black flies than are others and indicate that these differences may be accounted for by individual production rates of breath CO2.

Identification of juvenile hormone III bisepoxide (JHB 3), juvenile hormone III and methyl farnesoate secreted by the corpus allatum of Phormia regina (Meigen), in vitro and function of JHB 3 either applied alone or as a part of a juvenoid blend

The corpus allatum of both male and female adult black blow fly, Phormia regina (Meigen), produced radioisotope-labeled juvenoids when incubated in vitro following a radiochemical assay. The production of juvenoids was greatly enhanced after the flies were fed a protein (liver) meal. These juvenoids were determined as juvenile hormone III bisepoxide (JHB 3), juvenile hormone III (JH III) and methyl farnesoate (MF). These three juvenoids were separated and identified by chromatography (TLC and HPLC) using synthesized standards, respectively. Then, JHB 3 standard and the biological samples of JHB 3 were further identified with PCI GS-MS using published spectrum data. Bioassay of synthetic JHB 3 revealed its ability to restore oögenesis in allatectomized, liver-fed female blow flies. The ability of these three juvenoids and their blends to restore oögenesis was the rational juvenoid blend (i.e. a mixture of JHB 3, JH III and MF according to the percent ratio secreted by the isolated CA of liver-fed females) > JHB 3 = JH III > MF = the irrational juvenoid blend (i.e. a mixture of an arbitrary ratio of JHB 3, JH III and MF). This is the first evidence suggesting that multiple juvenoid production and reception may be a normal event for insects.

Oviposition aggregation pheromone in the Simulium damnosum complex

Communal oviposition by the Simulium damnosum complex of Afrotropical blackflies (Diptera: Simuliidae) was investigated under controlled laboratory conditions, using wild-caught flies in Sierra Leone. Volatile compounds emitted by Simulium eggs were trapped using a closed collection system, and their attractiveness to gravid flies was tested in a two-choice behavioural bioassay. Significantly more female blackflies oviposited on substrates baited with freshly laid eggs (100% chose the baited substrate), or with the volatiles collected from freshly laid eggs (85% chose the baited substrate), in preference to the relevant control substrates. Substrates baited with volatiles from 12-h-old eggs were not significantly more attractive than controls (only 31% chose the baited substrates; P = 0.33). Gas chromatographic analysis of the egg volatiles consistently showed two peaks emanating from fresh eggs, but significantly lower amounts from 12-h-old eggs (P < 0.05). A novel system for collecting the volatiles from this and other blackfly species, as they laid eggs on a substrate in flowing water, is described. Volatiles collected using this method showed identical gas chromatographic profiles to those of fresh eggs alone, indicating that the flies themselves produced no other volatile chemical signals during oviposition. Evidently communal oviposition by S. damnosum s.l. was mediated by a pheromone emanating from fresh eggs. The role of pheromone-mediated egg aggregation in blackfly ecology is discussed, and its possible manipulation is considered.

Structure and function of the cibarial armature in Simuliidae

Cibarial armature morphology in adult female blackflies (Diptera: Simuliidae) is described using scanning electron microscopy. Three distinct types of armature are recognized, comprising those with teeth, e.g. Simulium ochraceum, S. ornatum, S. veracruzanum and S. vorax, those with spicules, e.g. Austrosimulium bancrofti, S. damnosum, S. exiguum, S. metallicum and S. neavei; and those lacking these projections, e.g. Prosimulium rufipes and S. lineatum. Whereas the armature is poorly developed in vectors of human onchocerciasis such as S. damnosum, S. exiguum, S. metallicum and S. neavei, the well-developed armature in S. ochraceum, S. veracruzanum and S. vorax does not prevent these species becoming infected with Onchocerca spp. (Nematoda: Onchocercidae). Hence the armature is not primarily a mechanism to counteract microfilaria superinfection. Since cibarial armatures are more developed in the haematophagous females than in the males of certain Families of flies, e.g. Ceratopogonidae, Culicidae, Phlebotominae and Simuliidae in the sub-order Nematocera, evidently the armature has evolved in response to the blood-feeding habit. As the suction of imbibed blood by the cibarial pump may require a valve mechanism to prevent back-flow, it is suggested that the armature is primarily for this purpose. Secondarily, the cibarial armature presents a damaging barrier against ingested microfilariae.

Factors influencing nuisance blackfly (Diptera: Simuliidae) activity in the Scottish Highlands

The nuisance activity of blackflies (Diptera: Simuliidae) was investigated in several habitats on Speyside, near Kincraig (57 degrees 08'N, 3 degrees 56'W), Invernesshire, in central Scotland during May-October 1987-89. The main blackfly species caught landing/biting on humans were Simulium reptans, S. argyreatum, S. variegetum and the S. tuberosum complex, in order of prevalence. Blackfly biting activity occurred from mid-May to mid-September. Numbers of female blackflies attracted to volunteers were correlated with their body posture, habitat and the season. Overall, Simulium activity was greatest in mixed birch/juniper forest, least in spruce plantations and at intermediate levels on pasture, moorland and in Scots Pine forest. Compared with an adult, a child experienced twice as many Simulium bites per hour (12.2 v 6.3) in the birch forest. On the child's body, 69% of blackflies landed on the head, neck and back, whereas the majority landed on the legs (48%) and arms (28%) of adults. Bending over, especially during the exertion of gardening, was more attractive to anthropophilic blackflies than standing or kneeling.

Copulatory Apparatus and Deposition of Male Accessory Secretion in Phormia regina (Diptera: Calliphoridae)

A component of the male accessory gland secretion, injected during mating, was responsible for onset of refractory behavior in the female black blow fly, Phormia regina (Meigen). Immediately after mating, two components of the semen were located separately in the bursa copulatrix: sperm were found anteriorly near the opening to the spermathecal ducts, and accessory secretion distended the posterior of the bursa. The accessory secretion disappeared within a day of mating, presumably entering the hemolymph from where it was detected. Scanning electron microscopy of the intromittent organ and phallosome, and reconstruction of the female reproductive tract from serial sections revealed complementary structures in the male and female. A pair of conspicuous flaps on the phallosome clasp a cuticular tonsil in the female, sealing the accessory secretion in the bursa during copulation. Accessory secretion is released at the tip of the phallosome and is redirected to a more proximal release site on the phallosome via a pair of hitherto undescribed accessory ducts, resulting in its deposition posteriorly in the bursa lumen.

Entomological evaluation of ivermectin mass treatment against onchocerciasis

To evaluate the impact of onchocerciasis control programmes on the annual transmission potential, it is customary to monitor the man-biting rate of vector Siniuliuin (Diptera: Simuliidae) and the number of Oizclzocerca (Nematoda: Onchocercidae) infective larvae found in them (Walsh et al., 1978). In addition to the transmission of Onchocerca volvulus (Leuckart) causing human onchocerciasis, Simuliunz females may also be the vectors of various other species of Onchocerca which parasitize wild or domestic animals. The larvae of other Oizclzocerca spp. are difficult to distinguish from those of O. volvulus. Therefore, using routine methods in the field, it is not easy to determine the efficiency of a mass host chemotherapy campaign targeting only one Oizclzocerca species. To evaluate the impact of ivermectin on natural transmission of O. volvulus, we tried to improve the usual entomological index of infection by measuring lengths of Onchocerca larvae found in female blackflies (Simuliidae) collected on human bait. Annual mass treatment with ivermectin has been carried out since 1987 in the Vina Valley, a savanna region of North Cameroon, involving about 20,000 people (Prod'hon et al., 1991). Ivermectin was given to eligible people at the target dosage of 150 kg per kilogram of body weight. Coverage with the treatment was higher than 60% (Prod'hon et al., 1991). According to Traore-Lamizana & Lemasson (1987), the principal vectors of onchocerciasis in Vina valley are Simulium damnosunz Theobald sensu stricto and Xsirbaizurn (Vajime & Dunbar). Entomological surveys of man-biting Simuliidae were performed 1 month before treatment; then done again during the 2 months following the campaign. Simiilium females were dissected and larvae of Onchocerca were observed and counted as described by Philippon (1977). All third stage larvae found in the blackfly head, considered to be infective larvae, were stretched by heat exposure, then measured using an eye-piece micrometer. Infective larvae were recorded per 1000 parous blackflies. Infective larva lengths were repre-