Genetics of hybridization of Glossina swynnertoni with Glossina morsitans morsitans and Glossina morsitans centralis

Abstract Reciprocal crosses were performed with Glossina swynnertoni and Glossina morsitans morsitans and with G. swynnertoni and Glossina morsitans centralis , using strains that carried marker genes in all three linkage groups. Glossina swynnertoni males can inseminate, but not fertilize, G.m.morsitans; all other crosses produced some fertile females. Hybridization did not cause sex ratio distortion among F_{ flies. Most F and backcross females were fertile, but all F, males were sterile. Sterility among backcross males was also high (99% in BXj, 85% in Bx₂, and about 50% in Bx₃ to Bx₅). Chromosome transmission by hybrid females usually conformed to Mendelian expectations, but genetic recombination was lower than observed in G.m.morsitans. The reduction in fertility among backcross females was not associated with heterozygosity in any linkage group. Sterility among hybrid and backcross males was associated with heterozygosity of sex chromosomes and probably autosomes. The results support the systematic placement of G.swynnertoni closer to G.mxentralis than to G.m.morsitans.     

Rhythmic pulses of haemolymph pressure associated with parturition and ovulation in the tsetse fly,Glossina morsitans

Abstract Expulsion of the tsetse larva from the uterus of the female is preceded by 1–2 h of rhythmic pulses of haemolymph pressure that can be detected using a barographic technique. At first baseline pressure is maintained and all pulses are positive in relation to baseline. Then, about 1 h before parturition, baseline pressure increases, pulse intensity increases, and the pulses become both positive and negative in relation to baseline. Each pulse correlates with ‘bobbing’ action of the female's proboscis, the only external indication of this internal activity. A single large pressure pulse is observed at parturition, and thereafter the pressure level returns to the original baseline and pulsing action ceases. Around the presumptive time of ovulation, 1–2 h after parturition, another series of pressure pulses is observed. The pulses are the likely consequence of coordinated waves of muscular contraction that are essential preparation for successful parturition and ovulation.     

Insect immunity: Inducible antibacterial activity in Drosophila

Adult Drosophila melanogaster were found to produce an inducible antibacterial activity as a response to an injection (vaccination) of the non-pathogenic bacterium, Enterobacter cloacae. Vaccinated flies showed an increased survival time after a second injection with an insect-pathogenic bacterium, Pseudomonas aeruginosa. This immune response was blocked by cycloheximide, an inhibitor of eukaryotic protein synthesis, a fact which indicates de novo synthesis of the antibacterial factor operating in vivo. Electrophoretic mobility at pH 4 in combination with antibacterial assay and immunological cross reactivity was used to demonstrate attacin-like factors in hemolymph and cecropin-like factors in cell-free extracts of whole flies. Extract of flies also contained lysozyme but this enzyme could not be induced. Some active material is pre-existing and can be released by treatment of whole flies with ultrasound (sonication) or microwaves. Compared to wild-type flies, a mutant strain deficient in leucine aminopeptidase showed much higher values for antibacterial activity and lysozyme.     

Thermal effect of blood feeding in the telmophagous fly Glossina morsitans morsitans

During feeding on warm-blooded hosts, haematophagous insects are exposed to thermal stress due to the ingestion of a meal which temperature may highly exceed their own body temperature. In order to avoid overheating and its subsequent deleterious effects, these insects respond by setting up molecular protective mechanisms such as heat shock proteins synthesis or by using thermoregulative strategies. Moreover, the duration of contact with the host depends on the way of feeding displayed by the different species (either telmophagous or solenophagous) and thus also impacts their exposure to heat. Solenophagous insects feed directly on blood vessels and are relatively slow feeders while telmophagous insects by lacerating capillaries, facilitate their access to blood and thus feed more quickly. The aim of this work was to investigate to what extent strictly telmophagous insects such as tsetse flies are exposed to thermal stress during feeding and consequently to evaluate the impact of the feeding strategy on the exposition to overheating in haematophagous insects in general. Real time thermographic analysis during feeding revealed that the flies’ body significantly heat up quite homogeneously. At the end of feeding, however, a marked regional heterothermy occurs as a consequence of the alary muscles warm up that precedes take-off. Feeding strategies, either solenophagy or telmophagy, thus appear to have a great impact on both exposition to predation risks and to thermal stress.     

Insect haemolymph: Cooperation between humoral and cellular factors in Locusta migratoria

In Locusta migratoria, prophenoloxidase is present in the plasma and serum, but in reduced amounts relative to the haemocytes. This phenoloxidase activity cannot be induced by either heating or freezing and thawing and it is lost by heating at 70°C for 30 min. Both lipopolysaccharides and laminarin can elicit the prophenoloxidase activating system. These elicitors of prophenoloxidase activation are active in haemocyte lysate and in serum but never induce any phenoloxidase activity in plasma. In haemocyte lysate, the activating system is not heat resistant, and heating at 56°C for 30 min prior to incubation with laminarin or lipopolysaccharide precludes any phenoloxidase activity. Plasma contains a strong inhibitor of the prophenoloxidase activating system but serum does not. This inhibitor does not affect the phenoloxidase enzyme itself. The possible role of the activating system in immune recognition and the strategies evolved by parasites or pathogens to escape being recognized by their host are discussed.     

Octopamine distribution in the tsetse fly Glossina morsitans

Tissues of Glossina morsitans were assayed for octopamine using an enzymatic technique. Octopamine was detected at the highest concentration in the brain (7.06-7.99 ng mg-1 tissue protein) and thoracic ganglion (10.9-13.89 ng mg-1 tissue protein). Octopamine was present in haemolymph at a concentration of 1.0-1.27 X 10(-7) M. This was not found to vary when insects were flown or mechanically stressed. Nervous tissue, flight muscle and haemolymph showed a significant ability to metabolize octopamine. The greatest enzyme activity was present in the haemolymph.     

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.     

Responses of tsetse flies, Glossina morsitans morsitans and Glossina pallidipes, to baits of various size

Recent studies of Palpalis group tsetse [Glossina fuscipes fuscipes (Diptera: Glossinidae) in Kenya] suggest that small (0.25 × 0.25 m) insecticide-treated targets will be more cost-effective than the larger (≥1.0 × 1.0 m) designs currently used to control tsetse. Studies were undertaken in Zimbabwe to assess whether small targets are also more cost-effective for the Morsitans group tsetse, Glossina morsitans morsitans and Glossina pallidipes. Numbers of tsetse contacting targets of 0.25 × 0.25 m or 1.0 × 1.0 m, respectively, were estimated using arrangements of electrocuting grids which killed or stunned tsetse as they contacted the target. Catches of G. pallidipes and G. m. morsitans at small (0.25 × 0.25 m) targets were, respectively, ～1% and ～6% of catches at large (1.0 × 1.0 m) targets. Hence, the tsetse killed per unit area of target was greater for the larger than the smaller target, suggesting that small targets are not cost-effective for use against Morsitans group species. The results suggest that there is a fundamental difference in the host-orientated behaviour of Morsitans and Palpalis group tsetse and that the former are more responsive to host odours, whereas the latter seem highly responsive to visual stimuli.     

Horizontal transmission of mycotic infection in adult tsetse,Glossina morsitans morsitans

AdultGlossina morsitans morsitans exposed to wet conidia ofBeauveria bassiana andMetarhizium anisopliae suffered high mortalities ranging from 90 to 100% by 2 weeks post-exposure. Infected ♂ ♂ maintained in the same cages with non-infected ♀♀ throughout the experimental period transmitted the fungal infection to the ♀♀ resulting in mortalities of 65% withB. bassiana and 55% withM. anisopliae. Likewise, infected ♀♀ maintained together with non-infected ♂♂ transmitted the infection to the ♂♂ resulting in mortalities of 75% withB. bassiana and 45% withM. anisopliae. Female tsetse flies infected withB. bassiana andM. anisopliae and maintained in the same cages with non-infected ♀♀ also transmitted infection to the non-infected tsetse resulting in mortalities of 62% and 48% withB. bassiana andM. anisopliae respectively. Infected tsetse exposed to non-infected tsetse of the opposite sex for only 30 min were also able to transmit the fungal infection. Pupae produced by female tsetse infected withB. bassiana andM. anisopliae exhibited higher pupal mortality than those produced by non-infected ♀♀. However, pupae exposed directly to dry spores ofB. bassiana andM. anisopliae had no increase in pupal mortality but adults emerging from theB. bassiana-exposed pupae had markedly reduced longevity.      

Binding and action of cecropin and cecropin analogues: antibacterial peptides from insects

The mechanism of action of cecropin was studied by using liposomes as a model system. The bilayer was efficiently destroyed if the liposome net charge was zero or negative. Cecropin analogues with an impaired N-terminal helix had reduced membrane disrupting abilities that correlate with their lower antibacterial activity. The reduced bactericidal activity of the analogues was rationalized in terms of reduced binding to bacteria. The stoichiometry of cecropin killing of bacteria suggests that amounts of cecropin sufficient to form a monolayer strongly modify the bacterial membrane. Although some bacteria were resistant to cecropin they did bind large amounts in a non-productive manner. In contrast, mammalian erythrocytes achieve resistance by avoiding the binding of cecropin.     

Identification of major soluble salivary gland proteins in teneral Glossina morsitans morsitans

Salivary glands of tsetse flies (Diptera: Glossinidiae) contain molecules that are involved in preventing blood clotting during feeding as well as molecules thought to be intimately associated with trypanosome development and maturation. Here we present a protein microchemical analysis of the major soluble proteins of the salivary glands of Glossina morsitans morsitans, an important vector of African trypanosomes. Differential solubilization of salivary proteins was followed by reverse-phase, high-performance liquid chromatography (HPLC) and analysis of fractions by 1-D gel electrophoresis to reveal four major proteins. Each protein was subjected to amino acid microanalysis and N-terminal microsequencing. A protein chemical approach using high-resolution 2-D gel electrophoresis and mass spectrometry was also used to identify the salivary proteins. Matrix-assisted, laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry and quadrupole time-of-flight (Q-TOF) tandem mass spectrometry methods were used for peptide mass mapping and sequencing, respectively. Sequence information and peptide mass maps queried against the NCBI non-redundant database confirmed the identity of the first protein as tsetse salivary gland growth factor-1 (TSGF-1). Two proteins with no known function were identified as tsetse salivary gland protein 1 (Tsal 1) and tsetse salivary gland protein 2 (Tsal 2). The fourth protein was identified as Tsetse antigen-5 (TAg-5), which is a member of a large family of anti-haemostatic proteins. The results show that these four proteins are the most abundant soluble gene products present in salivary glands of teneral G. m. morsitans. We discuss the possible functions of these major proteins in cyclical transmission of African trypanosomes.     

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.     

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.