Critical thermal limits depend on methodological context

A full-factorial study of the effects of rates of temperature change and start temperatures was undertaken for both upper and lower critical thermal limits (CTLs) using the tsetse fly, Glossina pallidipes. Results show that rates of temperature change and start temperatures have highly significant effects on CTLs, although the duration of the experiment also has a major effect. Contrary to a widely held expectation, slower rates of temperature change (i.e. longer experimental duration) resulted in poorer thermal tolerance at both high and low temperatures. Thus, across treatments, a negative relationship existed between duration and upper CTL while a positive relationship existed between duration and lower CTL. Most importantly, for predicting tsetse distribution, G. pallidipes suffer loss of function at less severe temperatures under the most ecologically relevant experimental conditions for upper (0.06 degrees C min(-1); 35 degrees C start temperature) and lower CTL (0.06 degrees C min(-1); 24 degrees C start temperature). This suggests that the functional thermal range of G. pallidipes in the wild may be much narrower than previously suspected, approximately 20-40 degrees C, and highlights their sensitivity to even moderate temperature variation. These effects are explained by limited plasticity of CTLs in this species over short time scales. The results of the present study have broad implications for understanding temperature tolerance in these and other terrestrial arthropods.     

Tsetse Salivary Gland Hypertrophy Virus: Hope or Hindrance for Tsetse Control?

MANY SPECIES OF TSETSE FLIES (DIPTERA: Glossinidae) are infected with a virus that causes salivary gland hypertrophy (SGH), and flies with SGH symptoms have a reduced fecundity and fertility. The prevalence of SGH in wild tsetse populations is usually very low (0.2%-5%), but higher prevalence rates (15.2%) have been observed occasionally. The successful eradication of a Glossina austeni population from Unguja Island (Zanzibar) using an area-wide integrated pest management approach with a sterile insect technique (SIT) component (1994-1997) encouraged several African countries, including Ethiopia, to incorporate the SIT in their national tsetse control programs. A large facility to produce tsetse flies for SIT application in Ethiopia was inaugurated in 2007. To support this project, a Glossina pallidipes colony originating from Ethiopia was successfully established in 1996, but later up to 85% of adult flies displayed symptoms of SGH. As a result, the colony declined and became extinct by 2002. The difficulties experienced with the rearing of G. pallidipes, epitomized by the collapse of the G. pallidipes colony originating from Ethiopia, prompted the urgent need to develop management strategies for the salivary gland hypertrophy virus (SGHV) for this species. As a first step to identify suitable management strategies, the virus isolated from G. pallidipes (GpSGHV) was recently sequenced and research was initiated on virus transmission and pathology. Different approaches to prevent virus replication and its horizontal transmission during blood feeding have been proposed. These include the use of antiviral drugs such as acyclovir and valacyclovir added to the blood for feeding or the use of antibodies against SGHV virion proteins. In addition, preliminary attempts to silence the expression of an essential viral protein using RNA interference will be discussed.     

Monitoring tsetse fly populations. I. The intrinsic variability of trap catches of Glossina pallidipes at Nguruman, Kenya

During 1986 the tsetse fly Glossina pallidipes Austen was monitored daily at Nguruman, southwestern Kenya, using three unbaited biconical traps. This was done to investigate the nature and causes of daily variation in trap catches. The variability of the observed catches was compared to a model which includes the trapping probability and the stochastic variation in the sex-ratio. By comparing the catches of male and female flies we are able to establish the sampling distribution of the trap catches. In addition to seasonal changes in the trap catches, day-to-day variations are observed and these are considered greater than the variation arising from the stochastic nature of the sampling process. Recommendations are made in relation to sampling tsetse fly populations.     

Inertia in physiological traits: Embryonopsis halticella caterpillars (Yponomeutidae) across the Antarctic Polar Frontal Zone

Geographic variation is characteristic of many physiological traits at the population and species levels. However, several recent studies have suggested that population-level variation is either limited or that it is mostly a consequence of phenotypic plasticity. Here we show that there is considerable physiological inertia in cold hardiness, upper thermal tolerance limits and desiccation resistance in caterpillars of the sub-Antarctic moth Embryonopsis halticella Eaton, such that populations from two climatically different islands are physiologically very similar. Both populations are moderately chill tolerant, with no difference in the supercooling points of caterpillars (-17 to -20 degrees C). Within their host plants caterpillars of both populations freeze at substantially higher, and statistically equivalent temperatures (-9.5 to -11.5 degrees C). The populations also have similar upper lethal limits (38 degrees C), and survival times of dry conditions (6-170 h depending on mass). The previously inexplicably low freezing point of caterpillars at the climatically less severe Marion Island seems likely a consequence of physiological inertia given that the freezing point of caterpillars within their hosts is only a few degrees below absolute minima at the older, and colder, Heard Island. Lack of adaptive geographic variation in physiological traits has consequences for models of range limits, and highlights the importance of exploring phenotypic plasticity as a response to climatic variation.     

Size and mortality rates of Glossina pallidipes in the semi-arid zone of southwestern Kenya

Seasonal changes in the mean size of tsetse, Glossina pallidipes Austen, as indicated by wing vein length, were monitored during 1983-86 at Nguruman, southwestern Kenya. Changes in size of nulliparous females and wing fray category 1 males were shown to be correlated with the relative humidity 2 months before they were captured. Soil temperature when flies were in the pupal stage had much less effect. Size dependent mortality was demonstrated, with the mean size of flies emerging from pupae significantly less than that of field-caught flies. This mortality must occur at emergence, since there was no evidence of size-dependent mortality once the flies became available to the trap. Size was correlated with density-independent mortality acting on the parent population 2 months previously. It might therefore be possible to use size as an index of the intensity of such mortality. This could be useful when assessing the level of additional mortality required to suppress tsetse populations.     

New Polymorphic Microsatellites in Glossina pallidipes (Diptera: Glossinidae) and Their Cross-Amplification in Other Tsetse Fly Taxa

We report the development and characterization of three new microsatellite markers in the tsetse fly, Glossina pallidipes (Diptera: Glossinidae). Fifty-eight alleles were scored in 192 individuals representing six natural populations. Allelic diversity ranged from 9 to 28 alleles per locus (mean 19.3 +/- 5.5). Averaged across loci, observed heterozygosity was 0.581 +/- 0.209, and expected heterozygosity was 0.619 +/- 0.181. Cross-species amplifications of the G. pallidipes loci in other tsetse fly taxa are reported.     

Tsetse flies are attracted to the invasive plant Lantana camara

In tsetse both sexes feed exclusively on the blood of vertebrates for a few minutes every 2-3 days. Tsetse flies seek cover from high temperatures to conserve energy and plants provide shelter for tsetse in all the biotopes they occupy. Recently, tsetse have taken cover in plantations and under the invasive bush Lantana camara that has invaded large areas of the tsetse fly belt of Africa. Flies from such refugia are implicated in sleeping sickness epidemics. In a wind tunnel we show that both foliage and an extract of volatiles from foliage of L. camara attract three tsetse spp. from different habitats: Glossina fuscipes fuscipes (riverine), G. brevipalpis (sylvatic) and G. pallidipes (savannah). Gas chromatography analysis of volatiles extracted from leaves and flowers of L. camara coupled to electroantennograme recordings show that 1-octen-3-ol and beta-caryophyllene are the major chemostimuli for the antennal receptor cells of the three tsetse spp. studied. A binary mixture of these products attracted these flies in the wind tunnel. The gas chromatography linked electroantennograme analysis of the L. camara extracts also show that the antennal receptor cells of the three tsetse spp. respond similarly to groups of volatiles derived from the major biosynthetic and catabolic pathways of plants, i.e. to mono- and sesquiterpenes, to lipoxidation products and to aromatics. Mixtures of these plant volatiles also attracted tsetse in the wind tunnel. These findings show that tsetse flies have conserved a strong sensitivity to volatile secondary products of plants, underlining the fundamental role of vegetation in tsetse survival.     

Complexity of the cold acclimation response in Drosophila melanogaster

Insects can increase their resistance to cold stress when they are exposed to non-lethal conditions prior to the stress; these plastic responses are normally described only in terms of immediate effects on mortality. Here we examine in Drosophila melanogaster the short- and longer-term effects of different conditions on several measures of cold resistance, but particularly chill coma recovery. Short-term exposure to sublethal temperature (cold hardening) did not decrease chill coma recovery times even though it decreased mortality. Exposure to 12 degrees C for 2 days (acclimation) decreased chill coma recovery times for a range of stressful temperatures when flies were cultured at 25 degrees C, but did not usually affect recovery times when flies were cultured at 19 degrees C. In contrast, 2-day exposure to 12 degrees C decreased mortality regardless of rearing temperature. Rearing at 19 degrees C decreased mortality and chill coma recovery time relative to rearing at 25 degrees C. Acclimation increased the eclosion rate of eggs from stressed females, but did not affect development time or size of the offspring. These results indicate that plastic responses to cold in D. melanogaster are complex when resistance is scored in different ways, and that effects can extend across generations.     

Effects of photoperiodically induced reproductive diapause and cold hardening on the cold tolerance of Drosophila montana

Coping with seasonal and daily variation in environmental conditions requires that organisms are able to adjust their reproduction and stress tolerance according to environmental conditions. Females of Drosophila montana populations have adapted to survive over the dark and cold winters at high latitudes and altitudes by spending this season in photoperiodically controlled reproductive diapause and reproducing only in spring/summer. The present study showed that flies of a northern population of this species are quite tolerant of low temperatures and show high seasonal and short-term plasticity in this trait. Culturing the flies in short day length (nearly all females in reproductive diapause), as well as allowing the flies to get cold hardened before the cold treatment, increased the cold tolerance of both sexes both in chill coma recovery time test and in mortality assay. Chill coma recovery time test performed for the females of two additional D. montana populations cultured in a day length where about half of the females enter diapause, also showed that diapause can increase female cold tolerance even without a change in day length. Direct linkage between diapause and cold tolerance was found in only two strains representing a high-altitude population of the species, but the phenomenon will certainly be worth of studying in northern and southern populations of the species with larger data sets.     

Cuticular lipid mass and desiccation rates in Glossina pallidipes: interpopulation variation

Abstract.  Tsetse flies, Glossina pallidipes (Diptera: Glossinidae) are said to have strong dispersal tendencies. Gene flow among these populations is estimated to be the theoretical equivalent of no more than one or two reproducing flies per generation, thereby raising the hypothesis of local regimes of natural selection. Flies were sampled from four environmentally diverse locations in Kenya to determine whether populations are homogeneous in desiccation tolerance and cuticular lipids. Cuticular hydrocarbon fractions known to act as sex pheromones do not differ among populations, thereby eliminating sexual selection as an isolating mechanism. Cuticular lipid quantities vary among populations and are not correlated with prevailing temperatures, humidities, and normalized density vegetation indices. Females demonstrate a stronger correlation than males between cuticular lipid mass and body weight. Desiccation rates also vary among populations, but are not correlated with the amounts of cuticular lipid. Chemical analysis of cuticular hydrocarbons by gas chromatography–mass spectroscopy shows that one of the four populations has more 11,15- and 11,21-dimethyl-31 hydrocarbon on females. These results are discussed in the context of population differences and estimates of gene flow.     

Monitoring tsetse fly populations.:II. The effect of climate on trap catches of Glossina pallidipes

In Part I it was shown that the sampling distribution of trap catches of tsetse flies, Glossina pallidipes Austen, at Nguruman, Kenya, using unbaited biconical traps follows a Poisson distribution. In this paper we examine the effect of humidity and temperature on day-to-day and seasonal variations in the trap catches. It is shown that the seasonal variation is significantly correlated with maximum daily temperature, the catches increasing with temperature when the maximum temperature is below 34 degrees C and decreasing with temperature when it is above 34 degrees C. The correlation between trap catches and relative humidity is not as good as the correlation with the maximum temperature, and the two together do not improve the fit to the trap catches. The day-to-day variation is significantly greater than the intrinsic variation due to the stochastic nature of the sampling process and for some traps it is correlated with temperature and humidity. An autoregressive model gives a half-life for the decay of departures from the mean of about 1 day and it is suggested that this indicates the movement of flies in response to animal movement or to climatic factors other than temperature or humidity. After removing the temperature dependent part of the seasonal variation and the autoregressive component of the data, the male and female catches are still significantly correlated.     

Interpopulational variation in recovery time from chill coma along a geographic gradient: a study in the common woodlouse, Porcellio laevis

Extreme temperatures restrict the performance of terrestrial arthropods and variations in low temperatures on a latitudinal scale influence physiological variables. Recovery time from chill coma is a measure of cold tolerance and it is a good index of climatic adaptation. We tested differences in recovery time of the common woodlouse (Porcellio laevis) exposed to different thermal conditions. Individuals were sampled from four different populations in Chile, spanning a latitudinal range of approximately 10 degrees . Significant differences were found in recovery time among experimental temperatures and among populations, but no interaction between these factors. The results of recovery time in P. laevis showed a positive increment with annual mean minimum temperature, indicating that there is geographical variation in recovery time. While body mass presented interpopulational variation, this variation was not associated with thermal variables or latitude. Overall, our results agree with previous studies in the sense that recovery time from chill coma decreases towards high latitudes, and it is independent of taxa, continent and hemisphere.     

The flight and landing of tsetse (Glossina) in response to components of host odour in the field

ABSTRACT. Studies were conducted in Zimbabwe of the responses of Glossina morsitans morsitans Westwood and Glossina pallidipes Austen to various host odours using either arrangements of electrocuting nets or visual observations. Tsetse flying upwind in a plume of carbon dioxide, acetone and octenol turned downwind upon flying into a plume of acetone or octenol, but did not turn upon flying into a plume of carbon dioxide. They also turned in response to a transient decline in odour concentration. Tsetse landed on the ground in the vicinity of a source of natural odour or artificial odour containing carbon dioxide but not at sources of acetone or octenol only. The proportion of female G.pallidipes caught at a source of natural odour (37%) was significantly different from that caught at a source of synthetic odour (17%). Resting tsetse stimulated by natural odour took off sooner than non-stimulated flies and had a strong upwind bias in the direction of take off. Tsetse stimulated with artificial odour did not take off sooner than non-stimulated flies. It is suggested that there is an unidentified components) of ox odour that activates resting tsetse.     

The effects of carbon dioxide anesthesia and anoxia on rapid cold-hardening and chill coma recovery in Drosophila melanogaster

Carbon dioxide gas is used as an insect anesthetic in many laboratories, despite recent studies which have shown that CO(2) can alter behavior and fitness. We examine the effects of CO(2) and anoxia (N(2)) on cold tolerance, measuring the rapid cold-hardening (RCH) response and chill coma recovery in Drosophila melanogaster. Short exposures to CO(2) or N(2) do not significantly affect RCH, but 60 min of exposure negates RCH. Exposure to CO(2) anesthesia increases chill coma recovery time, but this effect disappears if the flies are given 90 min recovery in air before chill coma induction. Flies treated with N(2) show a similar pattern, but require significantly longer chill coma recovery times even after 90 min of recovery from anoxia. Our results suggest that CO(2) anesthesia is an acceptable way to manipulate flies before cold tolerance experiments (when using RCH or chill coma recovery as a measure), provided exposure duration is minimized and recovery is permitted before chill coma induction. However, we recommend that exposure to N(2) not be used as a method of anesthesia for chill coma studies.     

Comparison between pars intercerebralis neurosecretory cells of Glossina and those of other higher Diptera

Three types of the A-neurosecretory cells (NSCs) were found in the pars intercerebralis (PI) of the adult brain of four species of tsetse flies (Glossina palpalis, G. pallidipes, G. austeni and G. morsitans). The typical formula of their composition is: 8 A1-, 14 A2- and 4 A3-cells. After permanganate oxidation, the neurosecretory materials (NSMs) of Al- and A3-cells are rich in strong acid groups. The A2-NSM contains both strong and weak acid groups after this treatment; besides, it retains a weak affinity to acid dyes. In addition to the A-cells, the PI of Glossina species seems to include a number of NSCs of the type B.
The data obtained indicate a great similarity of the NSC composition in the PI of Glossina and other related higher dipterans.
In addition to the NSCs, giant neurons and particular cells with large cytoplasmic inclusions were also found in the PI of Glossina. The cells with large cytoplasmic inclusions correspond apparently to vacuolated cells previously observed in the brain of some flies. It is suggested that the considerable complication in reproductive biology which took place in tsetse flies had no effect on the composition of the NSCs in their PI when compared to that in related species with more ordinary cycles of reproduction.     

Less is more: restricted application of insecticide to cattle to improve the cost and efficacy of tsetse control

Studies were carried out in Zimbabwe of the responses of tsetse to cattle treated with deltamethrin applied to the parts of the body where most tsetse were shown to land. Large proportions of Glossina pallidipes Austen (Diptera: Glossinidae) landed on the belly ( approximately 25%) and legs ( approximately 70%), particularly the front legs ( approximately 50%). Substantial proportions of Glossina morsitans morsitans Westwood landed on the legs ( approximately 50%) and belly (25%), with the remainder landing on the torso, particularly the flanks ( approximately 15%). Studies were made of the knockdown rate of wild, female G. pallidipes exposed to cattle treated with a 1% pour-on or 0.005% suspension concentrate of deltamethrin applied to the (a) whole body, (b) belly and legs, (c) legs, (d) front legs, (e) middle and lower front legs, or (f) lower front legs. The restricted treatments used 20%, 10%, 5%, 2% or 1% of the active ingredient applied in the whole-body treatments. There was a marked seasonal effect on the performance of all treatments. With the whole-body treatment, the persistence period (knockdown > 50%) ranged from approximately 10 days during the hot, wet season (mean daily temperature > 30 degrees C) to approximately 20 days during the cool, dry season (< 22 degrees C). Restricting the application of insecticide reduced the seasonal persistence periods to approximately 10-15 days if only the legs and belly were treated, approximately 5-15 days if only the legs were treated and < 5 days for the more restricted treatments. The restricted application did not affect the landing distribution of tsetse or the duration of landing bouts (mean = 30 s). The results suggest that more cost-effective control of tsetse could be achieved by applying insecticide to the belly and legs of cattle at 2-week intervals, rather than using the current practice of treating the whole body of each animal at monthly intervals. This would cut the cost of insecticide by 40%, improve efficacy by 27% and reduce the threats to non-target organisms and the enzootic stability of tick-borne diseases.     

Effect of host packed cell volume on the bloodmeal size of male tsetse flies, Glossina pallidipes

aematin contents of engorged, male tsetse flies, Glossina pallidipes Austen, were compared with the packed cell volumes of oxen on which they had fed. Haematin contents icnreased with packed cell volume up to packed cell volumes of approximately 30%. Haematin contents appeared to level off or decline with further increase in packed cell volume. These results support a model of blood-feeding in tsetse flies in which the rate of blood consumption decreases as packed cell volume increases, because of increase in blood viscosity, and tsetse are unable to compensate for the decrease in consumption rate by feeding for a longer time. After allowing for the effects of packed cell volume, bloodmeal sizes of tsetse increased with ox body temperature.     

Use of fluorescent pigments for the automatic marking of tsetse flies in traps

A means of contaminating tsetse flies in the field with fluorescent pigment powders has been developed, using pigment in open-ended plastic chambers at the cage position on traps. Glossina pallidipes Austen and G.morsitans morsitans Westwood passed rapidly through the chambers, and on exit were contaminated with consistent doses of powder: about 90 micrograms/fly when powder was presented on the chamber roof and about 28 micrograms/fly when powder was presented on the chamber floor. The technique automatically marks tsetse flies with pigment, cheaply, simply and with the minimum imposition of stress and is expected to be particularly useful in ecological studies. Its potential for marking other biting flies is discussed.     

Behaviour of tsetse flies (Glossina) in host odour plumes in the field

ABSTRACT. In Zimbabwe, studies were made of the responses of Glossina pallidipes Austen and G.morsitans morsitans Westwood to artificial host odour using an incomplete ring of electrocuting nets. In a plume of synthetic host odour tsetse flew generally upwind, with 50–60% flying within 35^o of due upwind. More than 80% of tsetse flew at < 50 cm above ground level. Upon losing contact with odour they executed a reverse turn within about 2 m, and upon regaining contact they turned upwind. There were no clear differences in the responses of G.m.morsitans and G.pallidipes. Using electrocuting nets lying horizontally on the ground it was found that tsetse landed in the vicinity of the odour source, the propensity to land being greater for G.pallidipes than for G.m, morsitans , greater for immature than mature flies, and greater for males than females.     

Impact of Salivary Gland Hypertrophy Virus Infection on the Mating Success of Male Glossina pallidipes: Consequences for the Sterile Insect Technique

Many species of tsetse flies are infected by a virus (GpSGHV) that causes salivary gland hypertrophy (SGH). Female Glossina pallidipes (Austen) with SGH symptoms (SGH+) have reduced fecundity and SGH+ male G. pallidipes are unable to inseminate female flies. Consequently, G. pallidipes laboratory colonies with a high prevalence of SGH have been difficult to maintain and have collapsed on several occasions. To assess the potential impact of the release of SGH+ sterile male G. pallidipes on the efficacy of an integrated control programme with a sterile insect technique (SIT) component, we examined the mating efficiency and behaviour of male G. pallidipes in field cages in relation to SGH prevalence. The results showed in a field cage setting a significantly reduced mating frequency of 19% for a male G. pallidipes population with a high prevalence of SGH (83%) compared to 38% for a male population with a low prevalence of SGH (7%). Premating period and mating duration did not vary significantly with SGH status. A high percentage (>80%) of females that had mated with SGH+ males had empty spermathecae. The remating frequency of female G. pallidipes was very low irrespective of the SGH status of the males in the first mating. These results indicate that a high prevalence of SGH+ in G. pallidipes not only affects colony stability and performance but, in view of their reduced mating propensity and competitiveness, releasing SGH+ sterile male G. pallidipes will reduce the efficiency of a sterile male release programme.