The role of sex pheromones in initiation of copulatory behaviour by male tsetse flies, Glossina morsitans morsitans

ABSTRACT. Copulatory responses to a variety of decoy objects baited with synthetic 15,19,23-trimethylheptatriacontane were induced in adult male Glossina morsitans morsitans Westw., provided the object was of similar shape and size to a female fly. Chemical and configurational (shape and texture) stimuli were important in eliciting and maintaining copulatory behaviour of males. A dose/response relationship was obtained from 7-day-old test males, with an ED₉₅ of 2 μg using as decoys solvent-washed dead males baited with 15,19,23-trimethylheptatriacontane. The quantity of this compound in 5-day-old adult females is more than 4.0 μg; in males, only 0.6 μg. It is considered to have the characteristics of a true contact sex pheromone for the species, and does not act synergistically with other less stimulatory synthetic compounds. Gas chromatographic analysis of adult female cuticular paraffins of G.m.morsitans showed the presence of two large peaks between C₃₇ and C₃₈ which were virtually absent in males. Young adults of both sexes possessed a series of hydrocarbons below C₂₉ which disappeared during the first few days of adult life. The natural pheromone appears on the pharate adult female about 2 days before emergence from the puparium and is present throughout life. This suggests it is a component of the surface cuticular waxes. Responsiveness of males to pheromone-baited decoys increased to a maximum 3–4 days after emergence in regularly fed flies and only declined during the 24 h prior to death from starvation.     

The causes of mating abstention in male tsetse flies Glossina morsitans

ABSTRACT. The sexual response of a Glossina morsitans morsitans Westw. male towards a decoy consisting of a dead male dosed with synthetic sex pheromone, was depressed by the presence of the cuticular lipids of the decoy male. However, addition of a C24 paraffin, not normally found on or in the cuticle of male or female G.m.morsitans , to a decoy male from which cuticular lipids were removed had a similar effect. Addition of either the C24 paraffin or an extract of male cuticular lipids to decoy females which possess natural sex pheromone in their cuticles, again depressed the sexual responses of males towards the decoys. The n-hexane solvent used in the application of compounds to decoys, itself depressed responses towards natural pheromone on a dead female at volumes in excess of 100μl, presumably because it mixed the surface waxes which masked the pheromone. Occlusion experiments implied that sex pheromone receptors are present on both the tarsi and tibiae of the male. No evidence was found for the existence of a specific mating deterrent on or in the cuticle of male G.m. morsitans.     

The roles of vision and olfaction in mate location by males of the tsetse fly Glossina morsitans morsitans

The roles of visual and/or olfactory stimuli in eliciting mating responses from male Glossina morsitans morsitans Westwood were examined, using a system for automatically recording the number and duration of mating strikes made towards decoys, under controlled conditions. The results confirm that there is no olfactory component of the female sex recognition pheromone sensed by the male antennae, and the attraction of males to females appears to be visual. The absence of male-male mating strikes was the result of the absence of female sex-pheromone, rather than the presence of a repellent mating deterrent in the male cuticle. Experiments with coloured, artificial, sex-pheromone-dosed, cotton decoys showed that colour had only weak effects on attractiveness and number of encounters with decoys, and that no colour caused significant enhancement of mating responses over those shown to decoy females.     

Comparison of the infection rate of tsetse, Glossina morsitans morsitans, fed in vitro or in vivo

Studies were made of infection rates of trypanosomes in the tsetse fly Glossina morsitans morsitans Westwood (Diptera: Glossinidae) when maintained in vivo (rabbits) or in vitro on high quality, gamma-irradiated, sterile defibrinated bovine blood, obtained from the Entomology Unit of the International Atomic Energy Agency (IAEA). For both Trypanosoma congolense Broden and T. b. brucei Plimmer & Bradford, in vitro maintenance significantly reduced the proportion of flies that developed mature metacyclic trypanosome infections.     

Post-feed buzzing in the tsetse, Glossina morsitans morsitans, is an endothermic mechanism

ABSTRACT. Post-feed buzzing in Glossina morsitans morsitans Westw. causes a rise in thoracic temperature relative to the length of the buzz. As lift is proportional to the square of wing-beat frequency, which increases with temperature up to 32°C, buzzing results in an increase in the lift which the fly can produce. Heat generated by buzzing, in combination with the heat received from the host at the time of feeding, may well allow the fly to maximize lift generated in the immediate post-feeding period. Buzzing flies excrete excess water from the meal more rapidly than non-buzzing flies. It is argued that this is due to a rise in abdominal temperature. Maximized lift in the immediate post-feeding period and the rapid elimination of water from the very large blood meals taken by these flies may be expected to have strong selective advantages for the flies.     

Tsetse mating behaviour: effects of age and hunger in Glossina morsitans morsitans and G.pallidipes

ABSTRACT. The effects of age and hunger on the responses of male Glossina morsitans morsitans Westwood and G.pallidipes Austen to freeze-killed female decoys, were examined in the laboratory. In both species, activity, estimated as the total number of interactions between males and decoys, increased with both age and hunger. Interactions were divided into short-stay (<60 s) and long-stay, full copulatory responses. In both species, young, unfed males were significantly less likely to attempt to copulate with a decoy after encounter than were fed males. Among fed males the proportion of interactions that proceeded to full copulatory attempts did not change with increasing age, but decreased consistently with increasing hunger. At all ages and hunger levels, G.pallidipes were more active than G.m.morsitans. However, after encountering a decoy, G.pallidipes were less likely to attempt to copulate than G.m.morsitans. In both species the duration of copulatory attempts did not change with age, but declined with increasing hunger. Copulatory attempts by G.pallidipes were significantly shorter than those of G.m.morsitans. The results are discussed in relation to the behaviour of tsetse in response to control devices such as traps and targets.     

Lipophorin from the tsetse fly, Glossina morsitans morsitans.

1. Lipophorin was isolated from the haemolymph of adult tsetse fly, Glossina morsitans morsitans, by ultracentrifugation in a potassium bromide density gradient. 2. The tsetse fly lipophorin (Mr congruent to 600,000) has a density of congruent to 1.11 g/ml and consists of two apoproteins, apolipophorin-I (apoLp-I, Mr congruent to 250,000) and apolipophorin-II (apoLp-II, Mr congruent to 80,000), both of which are glycosylated as shown by staining with periodate-Schiff reagent. The protein complex is composed of 49% protein and 51% lipids. 3. The finding of lipophorin in tsetse fly haemolymph suggests that, although these flies primarily utilize proline for their energy needs, there is an active transport mechanism for the supply of lipid requirements.     

Activity patterns in pregnant tsetse flies, Glossina morsitans

ABSTRACT. Actograph measurements of the spontaneous flight activity of isolated female Glossina morsitans morsitans Westw. were made through their first two pregnancy cycles in constant conditions under LD 12:12. Activity fell from a mean of c. 20 units/day to near zero during the 24 h preceding the day on which larvipositionoccurred. Activity on the day of larviposition rose significantly, to c. 35 units/day, then fell back to the baseline 20 units on subsequent days. The circadian pattern of activity was changed markedly by larviposition: 48 h before it, activity in the morning peak of the usual V-shaped diel pattern was twice normal; over 70% of the flies then fed, and the evening arm of their V disappeared; 24 h before larviposition, most flies were totally inactive, the few that did fly performing only 1 or 2 units/day, and only in the first and last hours of the photophase; on the day of larviposition, the morning peak of the V was reduced by c. 70%, and activity peaked instead during the post-noon 2–3 h, before and during the act of parturition. Almost all larvipositions occurred between noon and lights-out. It is suggested that the c. 48 h of pre-larviposition quiescence may play a role in the fact that pregnant G. morsitans females often survive non-residual aerosol insecticide campaigns.     

Biosynthesis of contact sex pheromone in the female tsetse fly, Glossina morsitans morsitans Westwood

Injection of [2,3 ¹⁴C] sodium succinate into recently emerged, unfed females of Glossina morsitans morsitans resulted in incorporation of radiolabel mainly into surface cuticular alkanes. In vivo experiments with intact flies showed that the distribution of labelled alkanes depended on fly mobility, the legs of unrestrained flies possessing proportionately larger amounts of radioactive hydrocarbon material than those of flies whose legs were tied together with a silk suture. The heads of both restrained and unrestrained flies contained proportionately more material per unit surface area than did any other body part. However, ablation experiments and in vitro incubation showed that the most active incorporation of label into alkanes occurred in the abdomen and that all dorsal abdominal segments were equally active. The ventral abdomen also incorporated label into cuticular alkanes in vitro, but other body parts were apparently less able to do so. The sex pheromone of G. m. morsitans is a trimethyl-substituted alkane, the labelling of which appeared to be in proportion to the relative abundance of its methyl groups among those of the other alkanes of the cuticle following injection of either intact or legless flies. Hence it is proposed that sex pheromone is synthesized along with other cuticular alkanes mainly by cells closely associated with the abdominal cuticle of females and that it is spread over the external surface both by diffusion and by grooming which leads to accumulations of hydrocarbon material on the legs.     

Ultrastructural studies of certain aspects of the development of Trypanosoma congolense in Glossina morsitans morsitans

The course of Trypanosoma congolense infections in Glossina morsitans morsitans was followed by electron-microscopic examination of ultrathin sections of the guts and proboscises of infected flies. Guts dissected from flies 7 days after infection with culture procyclic forms of T. congolense had heavy trypanosome infections in the midgut involving both the endo- and ectoperitrophic spaces. Trypanosomes were also seen in the process of penetrating the fully formed peritrophic membrane in the central region of the midgut. By post infection day 21, trypanosomes had reached the proboscis of the fly and were found as clumps of epimastigote forms attached to the labrum by hemidesmosomes between their flagella and the chitinous lining of the food canal. Desmosome connections were observed between the flagella of adjacent epimastigotes. Flies examined at postinfection days 28 and 42 had, in addition to the attached forms in the labrum, free forms in the hypopharynx.     

Structural studies on the major milk gland protein of the tsetse fly, Glossina morsitans morsitans.

1. The major protein in the milk gland secretions of the tsetse fly, Glossina morsitans morsitans, was isolated by a combination of gel permeation chromatography and crystallization. 2. It has a native Mr approximately 47,000 and is composed of two identical polypeptide chains (Mr approximately 21,000) as determined by chemical cross-linking studies. The protein has no covalently-bound carbohydrates or lipids. Amino acid analysis of the protein revealed relatively high amounts of the aromatic amino acids, tyrosine (9.1 mol.%) and phenylalanine (8.5 mol.%). Immunoblotting experiments using antiserum against the protein revealed no cross-reactivity with any other milk proteins. 3. Quantitation of the protein during the pregnancy cycle showed that synthesis of the protein by the milk glands of adult female flies starts as the larva moults into second instar and rapidly declines as it matures into third instar. 4. It is proposed that the major milk gland protein could provide essential amino acids needed for the puparium formation.     

Influence of temperature changes on larviposition rhythm in the tsetse fly, Glossina morsitans

ABSTRACT. The hourly and quarter-hourly distribution of larval deposition by female Glossina morsitans Westw. was studied at various temperatures. At constant temperature and humidity, peak larviposition occurred 8–9 h after lights on. The effect of confining the flies to daily (LD 12:12 h) regimes of at least twenty-five consecutive cycles that included a sudden increase in temperature of c. 6°C for 2 h was a significant reduction in depositions during each period of increased temperature, and a significant increase before the heat impulse. When the temperature was allowed to fall rapidly, larviposition rate doubled in the following 15 min.     

Development of a system for sterilizing tsetse flies, Glossina spp., in the Held

Various autosterilizing systems were evaluated on natural populations of Glossina pallidipes Austen and G.morsitans morsitans Westwood in Zimbabwe. These involved a clear plastic tower of two or three chambers mounted at the cage position on a trap. Inside one chamber flies could be exposed to the vapour phase of the chemosterilant bisazir, P,P-bis (1-aziridinyl)-N-methylphosphinothioic amide. The system was designed to encourage flies to enter the sterilizing chamber, delaying their exit for sufficient time to expose them to a sterilizing dose. This was accomplished in the most effective system by restriction of the exit to one small hole (6 mm diameter) at the roof-side junction. The number of flies remaining in the chamber declined exponentially. The rate of exit was directly proportional to the density of flies in the sterilizing chamber and to the number of exit holes. The probability of a fly being in this chamber for at least 1 or 7 min (the times taken for female and male G.m.morsitans respectively to receive an ED50) was 0.84 and 0.67 respectively with one fly present. With sixteen flies, these probabilities were 0.76 and 0.18 respectively. Results suggest that it may be possible to develop a cheap, safe and efficient autosterilizer for use on tsetse traps.     

Cloning and expression of the yolk protein of the tsetse fly Glossina morsitans morsitans

Two major families of nutritional proteins exist in insects, namely the vitellogenins and the yolk proteins. While in other insects only vitellogenins are found, cyclorraphan flies only contain yolk proteins. Possible sites of yolk protein synthesis are the fat body and the follicle cells surrounding the oocyte. We report the cloning of the yolk protein of the tsetse fly Glossina morsitans morsitans, a species with adenotrophic viviparity. The tsetse fly yolk protein could be aligned with other dipteran yolk proteins and with some vertebrate lipases. In contrast to the situation in most fly species, only a single yolk protein gene was found in the tsetse fly. Northern blot analysis showed that only the ovarian follicle cells, and not the fat body represents the site of yolk protein synthesis.     

Juvenile hormone mimics as effective sterilants for the tsetse fly Glossina morsitans morsitans

The development of puparia of Glossina morsitans morsitans Westwood was disrupted by topical applications of the juvenile hormone mimics S-methoprene (the resolved enantiomer of 11-methoxy-3,7,11-trimethyl-2,4-dodecadienoic acid 1-methyl ester) (Zoecon), S21149 (propionaldoxime-0-4-phenoxyphenoxyethylether) (Sumitomo), or S31183 (2-[1-methyl-2-(4-phenoxyphenoxy)ethoxy]pyridine) (Sumitomo) dissolved in acetone. Puparia so treated during the first 4 days of life suffered developmental abnormalities, the severity of which were dose-dependent. Similarly, puparia produced by adult females treated with these compounds were abnormal. Dose-response data showed that effects were greatest with S31183 and least with S-methoprene. Abnormalities in the form of abdominal lesions and wing crumpling were typical of flies emerging from puparia produced by S-methoprene-treated females. However, arrested development at the red eye and pigmented seta stage within the puparium were typical of offspring of females treated with S21149 and S31183. A dose of 2 micrograms per female of S31183 was sufficient to prevent emergence of offspring produced for the rest of the life of the fly. The same dose resulted in partial recovery of females treated with S21149 some 18 days following treatment. Treatment with 2 micrograms S-methoprene did not suppress completely the production of normal offspring and recovery was complete some 27-35 days after treatment. Exposure of males to 20 micrograms S31183 did not impair their ability to inseminate females; transfer of material during copulation was sufficient to prevent the production of viable offspring by their mates.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cues used by the black fly,Simulium annulus,for attraction to the common loon (Gavia immer)

The parasitic relationship between a black fly, Simulium annulus, and the common loon (Gavia immer) has been considered one of the most exclusive relationships between any host species and a black fly species. To test the host specificity of this blood‐feeding insect, we made a series of bird decoy presentations to black flies on loon‐inhabited lakes in northern Wisconsin, U.S.A. To examine the importance of chemical and visual cues for black fly detection of and attraction to hosts, we made decoy presentations with and without chemical cues. Flies attracted to the decoys were collected, identified to species, and quantified. Results showed that S. annulus had a strong preference for common loon visual and chemical cues, although visual cues from Canada geese (Branta canadensis) and mallards (Anas platyrynchos) did attract some flies in significantly smaller numbers.     

Ultrastructural Studies of Certain Aspects of the Development of Trypanosoma congolense in Glossina morsitans morsitans*

SYNOPSIS The course of Trypanosoma congolense infections in Glossina morsitans morsitans was followed by electron-microscopic examination of ultrathin sections of the guts and proboscises of infected flies. Guts dissected from flies 7 days after infection with culture procyclic forms of T. congolense had heavy trypanosome infections in the midgut involving both the endo- and ectoperitrophic spaces. Trypanosomes were also seen in the process of penetrating the fully formed peritrophic membrane in the central region of the midgut. By post infection day 21, trypanosomes had reached the proboscis of the fly and were found as clumps of epimastigote forms attached to the labrum by hemidesmosomes between their flagella and the chitinous lining of the food canal. Desmosome connections were observed between the flagella of adjacent epimastigotes. Flies examined at postinfection days 28 and 42 had, in addition to the attached forms in the labrum, free forms in the hypopharynx.     

Determining the age of adult male and female Glossina morsitans morsitans using a new technique

Abstract. 1. Pteridines accumulated linearly with age up to at least 63 days in male and 140 days in female Glossina morsitans morsitans Westwood kept under laboratory conditions. Time is the major factor influencing total pteridine accumulation levels with the accumulation rate being modulated by temperature.
2. It is suggested that pteridine accumulation may be used to determine the age of male and female tsetse flies in the field.