Oviposition pheromone in the Simulium damnosum complex: biological activity of chemical fractions from gravid ovaries

Abstract. Communal oviposition in the Afrotropical blackfly species complex Simulium damnosum Theobald (Diptera: Simuliidae) is mediated by a pheromone emitted by freshly laid eggs. Previously, two compounds (designated peaks A and B) emanating from fresh eggs were shown to be associated with attractiveness to gravid blackflies in bioassay. The present study investigated the role of these compounds by testing the responses of wild-caught Simulium yahense in Ghana to fractionated hexane extracts of gravid ovaries prepared by gas chromatography (GC). Although the fractions were prepared from Sierra Leonean Simulium leonense , GC analysis of the emissions from fresh S.yahense eggs showed that the volatile blends of both species were similar. When tested in a two-choice bioassay, 66% of ovipositing blackflies chose the substrate baited with a mixture of the four fractions recombined. In a series of bioassays testing responses to the four individual fractions presented with a control in a multiple-choice arrangement, only fraction 3 (containing peaks A and B) attracted significantly more ovipositions than the other fractions and control. However, fraction 3 failed to elicit a significant response when presented as the sole attractant with a control in a two-choice bioassay. It was concluded that fraction 3 , though mainly responsible for mediating aggregated oviposition by S.yahense , was acting in tandem with additional cues, probably further chemicals, which remain to be isolated and characterized.     

Aggregated oviposition in the Simulium damnosum complex is mediated by eggs in a laboratory bioassay

Abstract. . The phenomenon of aggregated oviposition in blackflies (Diptera: Simuliidae) was investigated under controlled laboratory conditions, using wild-caught females of the Simulium damnosum complex in Sierra Leone. A method was developed for inducing Simulium females to lay eggs, and used as a bioassay to measure the responses of gravid S. damnosum s.l. to freshly laid eggs of the same species complex. In a series of two-choice tests, significantly more ovipositing flies chose substrates already containing eggs over control substrates ( P = 0.004). The time from introduction of flies into the oviposition system to the onset of egg-laying was significantly less when eggs were already present ( P = 0.049). Flies responded more quickly when more eggs were present and the relationship between egg-batch number and the time of this response was curvilinear ( P = 0.012). Ecological advantages and disadvantages of such aggregation behaviour and the possible role of semiochemicals in its mediation are discussed.     

Identification of Communal Oviposition Pheromones from the Black Fly Simulium vittatum

The suite of pheromones that promote communal oviposition by Simulium vittatum, a North American black fly species, was identified and characterized using gas chromatography-mass spectrometry, electrophysiological, and behavioral bioassays. Behavioral assays demonstrated that communal oviposition was induced by egg-derived compounds that were active at short range and whose effect was enhanced through direct contact. Three compounds (cis-9-tetradecen-1-ol, 1-pentadecene, and 1-tridecene) were identified in a non-polar solvent extract of freshly deposited S. vittatum eggs that were capable of inducing the oviposition response. Electroantennography demonstrated that two of these three compounds (1-pentadecene and 1-tridecene) actively stimulated antennal neurons. Identification of the oviposition pheromones of this family may be helpful in developing control measures for nuisance black flies and for medically-important species such as Simulium damnosum sensu lato.     

Oviposition response of Simulium reptans (Diptera: Simuliidae) to the presence of conspecific eggs

Abstract.

• 1 The oviposition behaviour of Simulium reptans L. is described from two sites on the River Spey, Scotland. Female aggregations were observed immediately downstream of oviposition sites and were composed mainly of gravid flies (range 60–80%0) together with smaller numbers that were either freshly bloodfed (range 2–17%) or infected with mermithid nematodes (range 0–30%).
• 2 The time from landing on the oviposition sites to the onset of oviposition was recorded. The time in the presence of greater than 1-day-old eggs did not significantly differ from sites with no eggs present. However, the presence of freshly laid or I-day-old eggs significantly shortened the time to onset of oviposition. The cues that elicit oviposition are unknown but it is speculated that they may involve a pheromone.

     

Molecular phytogeny and typing of blackflies (Diptera: Simuliidae) that serve as vectors of human or bovine onchocerciasis

Abstract. A subregion of the mitochondrial large subunit (16s) rRNA gene was amplified by polymerase chain reaction (PCR) from nine species of blackflies (Diptera: Simuliidae) which serve as natural or experimental vectors of human or bovine Onchocerca parasites. PCR products from each species of blackfly were tested by directed heteroduplex analysis (DHDA), and their genotypes established according to diagnostic banding patterns of the heteroduplex products. Three alleles of mitochondrial 16s rRNA were found to exist in members of the Simulium (Ewardsellum) damnosum sensu lato complex from West Africa, and two alleles were found in the Neotropical Simulium (Psilopelmia) ochraceum Walker complex and the Simulium (Simulium) metallicum Bellardi complex. Different single alleles were detected in Austrosimulium bancrofti , in English S.(S.)noelleri and in two North American laboratory vectors: Simulium (Psilozia) vittatum Zetterstedt and S.(S.)decorum Walker. Phylogenetic analysis of 16s sequences indicated that blackflies from West Africa and the Americas formed distinct clades. Neotropical onchocerciasis vectors were found to be more closely related to Nearctic and Palaearctic non-vector Simulium species than to the African vectors of onchocerciasis.     

Semiochemical mediation of oviposition by the phlebotomine sandfly Lutzomyia longipalpis

Abstract. Extracts of rabbit food, hay and rabbit faeces elicited a positive oviposition response from gravid female Lutzomyia longipalpis sandflies (Diptera: Phlebotominae). Combined extract of rabbit food and oviposition pheromone had a synergistic effect on sandfly egg-laying, greatly increasing the number of eggs laid and resulting in a highly targeted response. Individually tubed flies, exposed to the combined extract, were shown to be 3.5 times more likely to survive oviposition and laid 2.5 times more eggs than control flies. A laboratory oviposition trap baited with the combined extract was tested in a cage and caught 31 (62%) of 50 gravid L.longipalpis over a 72h period.     

Influence of Substrate Color on Oviposition Behavior,Egg Hatchability,and Substance of Egg Origin in the Mosquito Anopheles sinensis (Wiedemann) (Diptera: Culicidae)

Understanding the factors that influence the choice of substrate for oviposition by the malaria mosquito is critical to efforts directed to the management of the disease and vector control measures aimed at modifications of larval habitat. The influence of black and white substrates on Anopheles sinensis (Wiedemann) (Culicidae: Anophelinae) female oviposition behavior and egg-hatching rate and the presence of substances associated with egg laying were studied. Results from the no-choice tests showed that the number of eggs laid on black substrate was significantly greater than that laid on white substrate. Results from the dual-choice tests revealed that gravid females showed stronger preference for the black substrate than that for the white substrate. Furthermore, the egg-hatching rate on white substrate was significantly lower than that observed on black substrate. Results from the three-choice tests showed that substance of egg origin was associated with the black substrates (UBS) that were attractive for and stimulated oviposition. The results of this study suggest that there might be some compounds in the black substrates which play a positive role in the oviposition behavior of female mosquitoes and in the development of eggs and that eggs might produce and release active substances that attract females and stimulate oviposition. These results could be important as regards to the optimization of mosquitoes raised for experimental purposes and the control of malaria mosquitoes by altering the oviposition behavior of gravid females.     

Changes in egg colour, egg weight and oviposition rate with the number of eggs laid by wild females of the small heath butterfly, Coenonympha pamphilus

ABSTRACT. 1. The colour of eggs laid by individual Coenonympha pamphilus (L.) (Lep., Satyridae) females changed over their lifespan. The first laid eggs were green, but after having laid about 100 eggs, females laid only yellow eggs.
2. By following females in the field, or by capturing wild females, and noting the colour of their eggs, it was established that younger females showed higher oviposition rates and laid heavier eggs than older females.
3. Hence, wild C.pamphilus females are unable to sustain a constant egg production by extracting amino acids or any other nutrients from nectar. We hypothesize that this is a general phenomenon in nectar feeding butterflies.
4. The egg colour did not seem to match the colour of the substrate on which the female chose to deposit the egg.     

Do Golden Egg Bugs (Phyllomorpha laciniata: Heteroptera, Coreidae) Require Conspecifics for Oviposition?

Golden egg bug (Phyllomorpha laciniata) females lay eggs on the bodies of conspecifics of both sexes. We investigated to what extent reproduction depended on the availability of conspecifics as oviposition substrate and the acceptability of the host plant as an alternative oviposition substrate in the absence of conspecifics. Mated females were placed in experimental enclosures each containing a sprig of fresh host plant. Each experimental female was subjected to one of three treatments: isolated from conspecifics (solitary), paired with another female, or paired with a male. Solitary females laid a few eggs on the host plant but then stopped laying eggs, and solitary females laid significantly fewer eggs than those enclosed with another female or a male. Females enclosed with a male laid no more eggs than those enclosed with a female. When two previously isolated females were later enclosed together, they soon renewed oviposition. Females in nature contained significantly more oviducal eggs than did females that were enclosed with other females for a short period. Thus the availability of suitable conspecifics as oviposition substrate stimulates the deposition of mature eggs, and reproduction depends on the presence of conspecifics of either sex as oviposition substrate.     

Oviposition habits of a univoltine blackfly, Prosimulium kiotoense (Diptera: Simuliidae), in Kyushu, Japan

Oviposition habits of a univoltine mammalophilic blackfly, Prosimulium kiotoense Shiraki, were investigated at a stream in Kyushu Island, Japan. The flies oviposited on bryophyte mosses growing on river bank rock surfaces. The eggs were laid singly, but large irregular egg masses were often formed because many females oviposited within the same small areas. Wet sites with dense bryophyte cover at heights between 0 and 15 cm above water level were selected for oviposition. Oviposition activity was observed in the latter half of April. P. kioteonse females came to the site to lay eggs after 11.00 hours in the morning, when air temperature rose to about 15 degrees C. Peak ovipositional activity occurred between 12.00 and 14.00 hours.     

Chemostimuli implicated in selection of oviposition substrates by the stable fly Stomoxys calcitrans

Horse and cow dung were tested as substrates for oviposition by the stable fly Stomoxys calcitrans (L) (Diptera: Muscidae) in laboratory cages. Odour alone from either horse or cow dung was sufficient to attract flies for oviposition. This was confirmed in wind tunnel experiments, where both horse and cow dung were shown to attract gravid stable flies. However, when S. calcitrans was offered a choice between these two oviposition substrates, flies always chose horse dung over cow dung, both when allowed to contact the substrates and when relying on dung odour alone. Analyses of volatile compounds emanating from horse and cow dung by gas chromatography linked antennogram recordings from S. calcitrans antennae revealed no differences in the chemostimuli released from the two substrates. The predominant chemostimulant compounds in both substrates were carboxylic acids (butanoic acid), alcohols (oct-1-en-3-ol), aldehydes (decanal), ketones (octan-3-one), phenols (p-cresol), indoles (skatole), terpenes (beta-caryophyllene) and sulphides (dimethyl trisulphide). Higher levels (20-40 p.p.m.) of carbon dioxide were recorded over horse dung compared with cow dung, a factor that may contribute to the preference exhibited by S. calcitrans for this substrate for oviposition.     

The effects of the timing of exposure to cadmium on the oviposition behavior of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Animals can greatly increase their fitness by choosing oviposition sites free from damaging substances such as the heavy metal cadmium (Cd). Previous research has shown that Drosophila melanogaster reared on uncontaminated media lay fewer eggs on substrates containing Cd. In this study, we examined the effects of prior exposure to Cd on oviposition site preference. We tested flies that had been exposed to Cd continuously from egg to adult, only during pre-adult stages, or for the 4 days of adulthood prior to testing. We found that flies avoid laying eggs on Cd-contaminated medium and also that flies exposed to Cd as adults laid significantly fewer eggs than controls never exposed to Cd. We did not observe consistent changes in oviposition as a result of pre-adult exposure alone. Our results suggest that the presence of cadmium in the environment, even at low doses and for short periods of time during adulthood, can harm the fitness of Drosophila melanogaster.     

Longitudinal evaluation of repellent activity of Ocimum gratissimum (Labiatae) volatile oil against Simulium damnosum

To determine the repellent activity of Ocimum gratissimum volatile oil against Simulium damnosum (blackflies), a 12 month (January-December 2003) field study was conducted in three onchocerciasis endemic communities (Idomido, Obio camp, and Ikot Adaha) in Ini Local Government Area of Akwa Ibom State, Nigeria. The result revealed that topical application of 20% (v/v) concentration of the oil with liquid paraffin as a base, reduced the biting rate of S. damnosum by 90.2, 81.6, and 79.7%, in Idomido, Obiocamp, and Ikot Adaha respectively. The oil gave protection against the bite of S. damnosum for at least 3 h. A total of 710 adults S. damnosum were caught by individuals treated with Ocimum oil, as against 4296 caught by the control group. When the flies caught by the treated individuals were dissected none of them was infected with microfilariae of Onchocerca volvulus. Human-vector contact and onchocerciasis transmission could be reduced by the topical application of the volatile oil during the peak biting periods of the vector     

Isolation of oviposition pheromone from the eggs of the sandfly Lutzomyia longipalpis

Abstract. Semiochemical components of eggs of the sandfly Lutzomyia longipalpis (Diptera: Psychodidae) were separated by high performance liquid chromatography. HPLC fractions were examined quantitatively and qualitatively by gas chromatography (GC). A bioassay was used to determine the oviposition attraction of gravid L. longipalpis to each of the fractions separately and a peak responsible for the semiochemical activity was identified. Gravid flies were placed in individual oviposition tubes to determine if the peak of interest was an oviposition stimulant. The active semiochemical fraction attracted gravid flies for oviposition. Furthermore, egg laying was enhanced: gravid flies exposed to the pheromone oviposited earlier and laid more eggs than control flies. GC analysis indicated that 1200 eggs (2 days old) gave a yield of 12.75 ug of active pheromone. This fraction had similar HPLC and GC retention times to caryophyllene oxide, suggesting comparable polarity and molecular weight.     

Allozyme genotypes of Drosophila buzzatii: feeding and oviposition preferences for microbial species, and habitat selection

Mature, mated female D. buzzatii were given a choice of nine microbial communities actively growing on cactus homogenate in laboratory population cages, and tests were made to determine if flies of different genotypes (for seven allozyme loci) chose different microorganism species for either feeding or oviposition. Variation in feeding preferences was determined from assays of electrophoretic genotypes and the ingested microorganism species of individual flies. Oviposition preference variation was analyzed indirectly by assaying the genotypes of individuals raised from eggs laid on different microorganisms. No significant evidence was found for differences in feeding preferences among adults of different genotypes. For oviposition preferences, there were significant microorganism-genotype associations for each of seven polymorphic loci. Analyses of the total electrophoretic genotype, rather than of individual loci, showed that the genotypes of eggs laid on the same microorganism species were more similar than those laid on different species. That is, females of different genotypes show habitat selection for oviposition sites, which would facilitate the maintenance of genetic polymorphisms.     

Oviposition of resistant and susceptible strains of Drosophila melanogaster in the presence of deltamethrin

Oviposition of three strains of Drosophila melanogaster in the presence of deltamethrin was observed. These strains had different levels of physiological susceptibility to deltamethrin. Two-choice tests were conducted with couples of flies in petri-dish arenas containing two oviposition dishes. On the first day of the experiment, females were given a choice between a treated oviposition dish and an untreated control dish. On the second day of the experiment, two control oviposition dishes were given to females. Although individual females showed a tendency to aggregate their eggs in one of the dishes, control experiments demonstrated an overall equal distribution of eggs between the dishes. When one of the two oviposition dishes in the arena was treated with deltamethrin, the percentage of females ovipositing and the mean number of eggs laid by females were reduced, compared with control arenas. Females avoided the treated oviposition dish and laid significantly more eggs on the control dish. Furthermore, when the deltamethrin concentration was increased on the first day, female flies postponed their oviposition and laid significantly more eggs on the second day. The resistant strain, SR, demonstrated the same capacity to select the untreated site for oviposition as the susceptible strain, but it showed a smaller oviposition reduction and egg retention. The relationship between physiological and behavioural susceptibility to deltamethrin is discussed.     

Oviposition preferences and antennal size in carrion flies

Carrion-feeding flies use odours emanating from the decomposing corpse as cues for oviposition and are described as generalists because the larvae feed on the corpses of diverse species. Whereas several features of the corpse may influence the oviposition choices of these flies, it is not known whether there is a preference for a particular species of corpse. We provided carrion flies with ovipositional (and feeding) choices in a field experiment, in which various odour sources were presented simultaneously. We found novel evidence of broadly consistent choices of carrion by flies from four families. Traps baited with decaying fish flesh captured the greatest number of individuals, whereas traps baited with decaying pig liver typically attracted the least. We also asked whether individuals captured in the various baits vary in antennal size, perhaps reflecting different capacities for odour detection. There was a trend for individuals of Lucilia sericata Meigen (Diptera: Calliphoridae) and the platystomatid collected from the traps baited with pig liver to have significantly larger antennae, whereas individuals of Muscina stabulans (Fallen) (Diptera: Muscidae) captured in traps baited with marine fish flesh had relatively longer antennae for their body size. Our data reveal a more nuanced pattern of oviposition behaviour in these generalist carrion flies, which may reflect differences in their preference of carrion with different nutrients, and in their capacity to detect particular odours.     

DNA probes for the identification of members of the Simulium damnosum complex (Diptera: Simuliidae)

Genomic libraries in plasmid have been constructed from various sibling species of blackflies of the Simulium damnosum complex from West Africa. Three cloned repetitive sequences, which show variation in copy number between sibling species, have been isolated. These clones can be used as probes for the dot-blot identification of larvae, pupae or adults into the three main West African subcomplexes, i.e. damnosum, squamosum and sanctipauli subcomplexes. The sequences also show some intraspecific variation in copy number.     

The thermal constant of the onchocerciasis vector Simulium damnosum s.l. in West Africa

The minimum water temperature for development (t(0)) and the thermal constant (K) for the development of immature stages of Simulium damnosum s.l. (Diptera: Simuliidae) in West Africa were estimated as 20.1 °C and 93 day-degrees, respectively, based on analyses of published data on development rates of eggs, larvae and pupae at different water temperatures (24.0 °C and 31.5 °C). Thus, at a constant water temperature of 30.0 °C (approximately 10 °C above t(0)), adult flies would emerge about 9 days after oviposition. Analysis of a dataset probably restricted to S. damnosum s.s., but for which the temperature for the egg stage varied, revealed a much lower t(0) (16.3 °C) and a much higher K (181 day-degrees), suggesting that the insects' thermal relations may be cytoform-specific. The results will aid control decisions and predictions of possible effects of climate change on sizes and geographic distributions of populations of onchocerciasis vectors in West Africa.     

Egg performance on an egg-carrying bug. Experiments in the field

Selection of oviposition sites has direct influence on female fitness. Differences in offspring survival among sites should favour females to select oviposition sites with the highest survival. Golden egg bug females ( Phyllomorpha laciniata; Coreidae, Heteroptera) use conspecifics as oviposition substrates. Most eggs are laid on the back of the bug but they can be found on all body parts. Females never carry their own eggs, and males commonly carry eggs received without copulation with the donor. We examined differences in egg survival relative to paternity to the egg, host sex and body size and attachment of an egg on a host's body in the field. We also studied which bugs received eggs. Egg loss was surprisingly common: 30–80% of recaptured individuals lost eggs during a six-day period. Unexpectedly, host's paternity to the eggs did not affect egg survival. Also, egg loss did not differ among hosts in respect to other parameters studied. Males received more eggs than females, but survival of the eggs was not better on a male's back than on a female's back. Since egg loss is common in the golden egg bug, we suggest that female oviposition strategy to lay eggs on conspecifics is not perfect. This is due to active behaviour of potential hosts (for example, rejected oviposition attempts) or lack of suitable oviposition substrates (conspecifics) in the field. Since eggs do not survive unless carried, females may be acting opportunistically, doing the best job possible by laying eggs on available conspecifics.