Disruption of the salivary gland muscle in tsetse, Glossina pallidipes Austen, as a result of salivary gland hypertrophy virus infection.

The secretory region of the salivary glands in Glossina pallidipes Austen (Diptera: Glossinidae) is characterized by an external muscle layer. Scanning electron microscopy and transmission electron microscopy investigations provide a detailed description of the longitudinal muscle fibres and a comparison of their structure when affected by salivary gland hypertrophy virus. The virus is responsible for hypertrophy of the salivary glands in symptomatic flies, specifically of the muscle fibres, the cytoarchitecture of which is completely altered. Although observations did not reveal viral particles in the muscle cells of either asymptomatic or symptomatic flies, muscle fibres were enlarged and detached from one another and their associated basement membrane only in symptomatic flies. A decrease in type IV collagen labelling in the basement membrane of the muscles in symptomatic flies is reported and is considered a potential cause of the salivary gland muscle alteration and, possibly, myopathy. The maintenance of an organized muscular layer is essential for the normal secretion of saliva and hence its pathology in symptomatic tsetse flies could affect the normal transmission of the trypanosome that develops inside the salivary gland epithelium. Therefore, a better understanding of the possible role of the virus is essential in order to elucidate its impact on salivary deployment in symptomatic flies.

Effect of wing length on the prevalence of trypanosomes in Glossina morsitans morsitans in eastern Zambia.

BACKGROUND: Tsetse flies (Diptera: Glossinidae) transmit trypanosomiasis (sleeping sickness in humans and nagana in livestock). Several studies have indicated that age, sex, site of capture, starvation and microbiome symbionts, among others, are important factors that influence trypanosome infection in tsetse flies. However, reasons for a higher infection rate in females than in males still largely remain unknown. Considering that tsetse species and sexes of larger body size are the most mobile and the most available to stationary baits, it was hypothesized in this study that the higher trypanosome prevalence in female than in male tsetse flies was a consequence of females being larger than males. METHODS: Black screen fly rounds and Epsilon traps were used to collect tsetse flies in eastern Zambia. Measurement of wing vein length and examination for presence of trypanosomes in the flies were carried out by microscopy. Principal component method was carried out to assess the potential of wing vein length as a predictor variable. The multilevel binary logistic regression method was applied on whole data, one-method data and one-sex data sets to evaluate the hypothesis. RESULTS: Data derived from a total of 2195 Glossina morsitans morsitans were evaluated (1491 males and 704 females). The wing length variable contributed the highest variance percentage (39.2%) to the first principal component. The variable showed significant influence on prevalence of trypanosomes when the analysis was applied on the whole data set, with the log odds for the prevalence of trypanosomes significantly increasing by 0.1 (P = 0.032), per unit increase in wing length. Females had higher trypanosome prevalence rates than males, though not always significant. Furthermore, moving from females to males, wing length significantly reduced by 0.2 (P < 0.0001). CONCLUSIONS: We conclude that wing length is an important predictor variable for trypanosome prevalence in Glossina morsitans morsitans and could partially explain the higher prevalence of trypanosomes in females than in males. However, reasonably representative population data are required for analysis-a serious challenge with the current tsetse sampling methods. Thus, analysis combining data from mobile and stationary methods that include both sexes' data could be useful to verify this hypothesis.

[Glossina feeding habits and diversity of species of trypanosomes in an active focus of human African trypanosomiasis in Gabon]. / Régime alimentaire des glossines et diversité des espèces de trypanosomes dans un foyer actif de trypanosomose humaine africaine au Gabon.

Feeding host is an important factor upon which depend the Glossina infection rate and the proportion of different species of trypanosome. Glossina feed both upon animals and humans. In order to identify species of trypanosomes present in the Komo-Mondah focus and to verify whether there is any relationship between the prevalence of sleeping sickness and the feeding habits of Glossina, we have carried out an entomological survey in this focus of Gabon. Flies were dissected and organs were analysed by PCR, while the origin of blood meals was determined by ELISA. Three species of trypanosomes were found: Trypanosoma congolense "forest type" (14/104; 13.46%), Trypanosoma vivax (11/104; 10.58%) and Trypanosoma brucei s.l. (65/104; 62.5%) with 13.46% (14/104) of mixed infections of T. brucei s.l. and T. congolense Glossina palpalis palpalis was caught in all biotopes investigated (91.85%) and was infected by all these species of trypanosomes. Glossina caliginea was not infected and Glossina fuscipes fuscipes was infected by T. brucei s.l. Tsetse flies feed more on animals than on humans in almost all villages, but there was no significant difference between the number of blood meals taken upon these two groups of vertebrates (Chi 2 = 7.43; p > 0.05). A negative correlation was found between the zoophylic/anthropophylic index and the prevalence of HAT. This result is insufficient to conclude that this index can be used as an indicator of the degree of prevalence of HAT. In contrary, the trypanosomian risk seems to be an appropriate indicator of the prevalence of HAT in an area. The identification of the reservoir hosts in this focus would be useful for a good understanding of the HAT epidemiology.

Wing length of tsetse caught by stationary and mobile sampling methods.

INTRODUCTION: A variety of techniques have been used to control tsetse with varying degrees of success. In a study on the population structure of Glossina fuscipes fuscipes that recovered after a previous vector control trial on two Kenyan islands, it was reported that the average fly size on the intervention islands was significantly smaller than on the none intervention islands and also compared to the size before the intervention. The conclusion was that vector control using tiny targets exerted size selection pressure on the population. The study recommended for further studies and suggested that this phenomenon could be among the reasons why targets used as a sole control method have rare reports of successful elimination of tsetse populations. Therefore, in this paper we report on a study of body size of tsetse flies caught in epsilon traps (as a stationary device) and black screen fly rounds (as a mobile trapping device). MATERIALS AND METHODS: The study was carried out in eastern Zambia to test the hypothesis that the body size (measured as wing length) of G. m. morsitans males or females, captured by epsilon traps and fly rounds is the same. RESULTS: A total of 1442 (489 females and 953 males) wing length measurements of G. m. morsitans were used in the analysis. It was established that tsetse flies caught by epsilon traps are on average larger than those caught by fly rounds. The likelihood of a large female or male fly being caught by traps, relative to a small one, significantly increased by 5.088 times (95% CI: 3.138-8.429) and by 2.563 times (95% CI: 1.584-4.148), respectively, p < 0.0001, compared with being caught by fly rounds. The hypothesis was rejected. CONCLUSION: This study showed that epsilon traps capture significantly larger G. m. morsitans than fly rounds do. Therefore, further research is recommended to verify i) whether the predilection of traps to capture larger flies has an effect on the process of tsetse elimination when targets are used e.g. targets may take longer to reach elimination than if the predilection was not there, ii) whether different results can be obtained on ecogeographic distribution of different sizes of the species if fly rounds are used for sampling instead of epsilon traps. The results from such studies could influence the strategies used in future control operations.

Experimental modifications imply a stimulatory function for male tsetse fly genitalia, supporting cryptic female choice theory.

One of the most sweeping of all patterns in morphological evolution is that animal genitalia tend to diverge more rapidly than do other structures. Abundant indirect evidence supports the cryptic female choice (CFC) explanation of this pattern, which supposes that male genitalia often function to court females during copulation; but direct experimental demonstrations of a stimulatory function have been lacking. In this study, we altered the form of two male genital structures that squeeze the female's abdomen rhythmically in Glossina pallidipes flies. As predicted by theory, this induced CFC against the male: ovulation and sperm storage decreased, while female remating increased. Further experiments showed that these effects were due to changes in tactile stimuli received by the female from the male's altered genitalia, and were not due to other possible changes in the males due to alteration of their genital form. Stimulation from male genital structures also induces females to permit copulation to occur. Together with previous studies of tsetse reproductive physiology, these data constitute the most complete experimental confirmation that sexual selection (probably by CFC) acts on the stimulatory properties of male genitalia.

Hearing in tsetse flies? Morphology and mechanics of a putative auditory organ.

Tympanal hearing organs are widely used by insects to detect sound pressure. Such ears are relatively uncommon in the order Diptera, having only been reported in two families thus far. This study describes the general anatomical organization and experimentally examines the mechanical resonant properties of an unusual membranous structure situated on the ventral prothorax of the tsetse fly, Glossina morsitans (Diptera: Glossinidae). Anatomically, the prosternal membrane is backed by an air filled chamber and attaches to a pair of sensory chordotonal organs. Mechanically, the membrane shows a broad resonance around 5.3-7.2 kHz. Unlike previously reported dipteran tympana, a directional response to sound was not found in G. morsitans. Collectively, the morphology, the resonant properties and acoustic sensitivity of the tsetse prothorax are consistent with those of the tympanal hearing organs in Ormia sp. and Emblemasoma sp. (Tachinidae and Sarcophagidae). The production of sound by several species of tsetse flies has been repeatedly documented. Yet, clear behavioural evidence for acoustic behaviour is sparse and inconclusive. Together with sound production, the presence of an ear-like structure raises the enticing possibility of auditory communication in tsetse flies and renews interest in the sensory biology of these medically important insects.

Using genetic and phenetic markers to assess population isolation within the southernmost tsetse fly belt in Africa.

The effective control of tsetse flies (Diptera; Glossinidae), the biological vectors of trypanosome parasites that cause human African trypanosomosis and African animal trypanosomosis throughout sub-Saharan Africa, is crucial for the development of productive livestock systems. The degree of genetic isolation of the targeted populations, which indicate reinvasion potential from uncontrolled areas, will be critical to establish a control strategy. Molecular and morphometrics markers were used to assess the degree of genetic isolation between seemingly fragmented populations of Glossina brevipalpis Newstead and Glossina austeni Newstead present in South Africa. These populations were also compared with flies from adjacent areas in Mozambique and Eswatini. For the molecular markers, deoxyribonucleic acid was extracted, a r16S2 Polymerase chain reaction (PCR) was performed and the PCR product sequenced. Nine landmarks were used for the morphometrics study as defined by vein intersections in the right wings of female flies. Generalised Procrustes analyses and regression on centroid size were used to determine the Cartesian coordinates for comparison between populations. Both methods indicated an absence of significant barriers to gene flow between the G. brevipalpis and G. austeni populations of South Africa and southern Mozambique. Sustainable control can only be achieved if implemented following an area-wide management approach against the entire G. brevipalpis and G. austeni populations of South Africa and southern Mozambique. Limited gene flow detected between the G. austeni population from Eswatini and that of South Africa or Mozambique may imply that these two populations are in the proses of becoming isolated.

Wing length and host location in tsetse (Glossina spp.): implications for control using stationary baits.

BACKGROUND: It has been suggested that attempts to eradicate populations of tsetse (Glossina spp.) using stationary targets might fail because smaller, less mobile individuals are unlikely to be killed by the targets. If true, tsetse caught in stationary traps should be larger than those from mobile baits, which require less mobility on the part of the flies. RESULTS: Sampling tsetse in the Zambezi Valley of Zimbabwe, we found that the number of tsetse caught from stationary traps, as a percent of total numbers from traps plus a mobile vehicle, was ~5% for male G. morsitans morsitans (mean wing length 5.830 mm; 95% CI: 5.800-5.859 mm) and ~10% for females (6.334 mm; 95% CI: 6.329-6.338 mm); for G. pallidipes the figures were ~50% for males (6.830 mm; 95% CI: 6.821-6.838 mm) and ~75% for females (7.303 mm, 95% CI: 7.302-7.305 mm). As expected, flies of the smaller species (and the smaller sex) were less likely to be captured using stationary, rather than mobile sampling devices. For flies of a given sex and species the situation was more complex. Multivariable analysis showed that, for females of both species, wing lengths changed with ovarian age and the month, year and method of capture. For G. pallidipes, there were statistically significant interactions between ovarian age and capture month, year and method. For G. m. morsitans, there was only a significant interaction between ovarian age and capture month. The effect of capture method was, however, small in absolute terms: for G. pallidipes and G. m. morsitans flies caught on the mobile vehicle had wings only 0.24 and 0.48% shorter, respectively, than flies caught in stationary traps. In summary, wing length in field samples of tsetse varies with ovarian age, capture month and year and, weakly, with capture method. Suggestions that a target-based operation against G. f. fuscipes in Kenya caused a shift towards a smaller, less mobile population of tsetse, unavailable to the targets, failed to account for factors other than capture method. CONCLUSIONS: The results are consistent with the successful use of targets to eradicate populations of tsetse in Zimbabwe. Until further, more nuanced, studies are conducted, it is premature to conclude that targets alone could not, similarly, be used to eradicate G. f. fuscipes populations in Kenya.

Molecular phylogenetics of tsetse flies (Diptera: Glossinidae) based on mitochondrial (COI, 16S, ND2) and nuclear ribosomal DNA sequences, with an emphasis on the palpalis group.

Relationships of 13 species of the genus Glossina (tsetse flies) were inferred from mitochondrial (cytochrome oxidase 1, NADH dehydrogenase 2 and 16S) and nuclear (internal transcribed spacer 1 of rDNA) sequences. The resulting phylogeny confirms the monophyly of the morphologically defined fusca, morsitans and palpalis subgenera. Genetic distances between palpalis and morsitans subspecies suggest that their status needs revision. In particular, cytochrome oxidase 1 sequences showed large geographical differences within G. palpalis palpalis, suggesting the existence of cryptic species within this subspecies. The morphology of palpalis group female genital plates was examined, and individuals were found varying outside the ranges specified by the standard identification keys, making definitive morphological classification impossible. A diagnostic PCR to distinguish G. palpalis palpalis, G. tachinoides and G. palpalis gambiensis based on length differences of internal transcribed spacer 1 sequences is presented.

Analysis of milk gland structure and function in Glossina morsitans: milk protein production, symbiont populations and fecundity.

A key process in the tsetse reproductive cycle is the transfer of essential nutrients and bacterial symbionts from mother to intrauterine offspring. The tissue mediating this transfer is the milk gland. This work focuses upon the localization and function of two milk proteins (milk gland protein (GmmMGP) and transferrin (GmmTsf)) and the tsetse endosymbionts (Sodalis and Wigglesworthia), in the context of milk gland physiology. Fluorescent in situ hybridization (FISH) and immunohistochemical analysis confirm that the milk gland secretory cells synthesize and secrete milk gland protein and transferrin. Knockdown of gmmmgp by double stranded RNA (dsRNA) mediated RNA interference results in reduction of tsetse fecundity, demonstrating its functional importance in larval nutrition and development. Bacterial species-specific in situ hybridizations of milk gland sections reveal large numbers of Sodalis and Wigglesworthia within the lumen of the milk gland. Sodalis is also localized within the cytoplasm of the secretory cells. Within the lumen, Wigglesworthia localize close to the channels leading to the milk storage reservoir of the milk gland secretory cells. We discuss the significance of the milk gland in larval nutrition and in transmission of symbiotic bacteria to developing offspring.

Population structuring of Glossina palpalis gambiensis (Diptera: Glossinidae) according to landscape fragmentation in the Mouhoun river, Burkina Faso.

The impact of landscape fragmentation due to human and climatic mediated factors on the structure of a population of Glossina palpalis gambiensis Vanderplank (Diptera: Glossinidae) was investigated in the Mouhoun river basin, Burkina Faso. Allele frequencies at five microsatellite loci, and metric properties based on 11 wing landmarks, were compared between four populations. The populations originated from the Mouhoun river and one of its tributaries. The average distance between samples was 72 km with the two most widely spaced populations being 216 km apart. The sampling points traversed an ecological cline in terms of rainfall and riverine forest ecotype, along a river enlarging from downstream to upstream and oriented south to north. Microsatellite DNA comparison demonstrated structuring between the populations, but not complete isolation, with an overall Fst = 0.012 (P < 0.001). Wing geometry revealed significant centroid size and shape differences between populations, especially between the two most distant populations. There was no significant correlation between gene flow and geographic distance at this scale, but there was a positive correlation in females between metric distances (wing shape differences) and geographic distances that might be attributed to the cline of environmental conditions. The impact of the fragmentation of riparian landscapes on tsetse population structure is discussed in the context of control campaigns currently promoted by Pan African Tsetse and Trypanosomosis Eradication Campaign.

The wing venation patterns to identify single tsetse flies.

This is the first study to explore the potential of various geometric morphometrics methods to help the morphological diagnostic of tsetse species, vectors of human and animal trypanosomiases in sub-Saharan Africa. We compared landmarks, semilandmarks and outlines techniques on male and female samples of species, and suggested adapted strategies according to the countries and their own Glossina fauna. We could compare up to 7 taxa belonging to the three main subgenera of the Glossina genus: Nemorhina (5 species), Glossina (1 species) and Austenina (1 species). Our sample included the major vectors of sleeping sickness: G. palpalis palpalis, G. p. gambiensis, G. fuscipes fuscipes and G. f. quanzensis, as well as two important vectors of African animal trypanosomoses: G. tachinoides and Glossina morsitans submorsitans. The average level of correct species recognition by the wing shape was satisfactory, and slightly higher for females than for males. The best scores of correct assignment, in both sexes, were obtained by the contour technique (96% of correct attribution in females, 92% in males), slightly higher than for semilandmarks (95% and 91%) or landmarks (94% and 89%) techniques. We made our images of wings freely available to be used as reference images (http://mome-clic.com), and we describe the conditions and the analytical steps to be followed to identify unknown specimens using external reference images. Under adequate conditions, such use of reference images obtained from a free access server could help species identification of new samples anywhere in Africa.

Morphometric diagnosis of Glossina palpalis (Diptera: Glossinidae) population structure in Ghana.

OBJECTIVE: This study aimed to identify isolated population(s) of Glossina palpalis in Ghana using geometric morphometrics to evaluate variations in wing-shape and size between populations of the fly from three regions. RESULTS: Wing shape of G. palpalis tsetse flies from the Northern, Western and Eastern Regions varied significantly between each other. Populations from the Northern and Western Regions varied the most (Mahalanobis Distance = 54.20). The least variation was noticed between populations from the Western and Eastern Regions (MD = 1.99). On morphospace, the Northern population clearly separated from the Eastern and Western populations both of which overlapped. Wing centroid size also significantly varied among populations. Reclassification scores were satisfactory reaching 100% for the Northern population. The Northern population of G. palpalis is possibly isolated from the Western and Eastern Region populations. Meanwhile, a panmictic relationship could be on-going between the Western and Eastern populations. We speculate that geographical distance and subspecific difference between populations are among factors responsible for observed pattern of wing shape variations among the studied populations. The implications of results regarding choice of control strategy and limitations of the study are discussed.

Microarchitecture of the tsetse fly proboscis.

BACKGROUND: Tsetse flies (genus Glossina) are large blood-sucking dipteran flies that are important as vectors of human and animal trypanosomiasis in sub-Saharan Africa. Tsetse anatomy has been well described, including detailed accounts of the functional anatomy of the proboscis for piercing host skin and sucking up blood. The proboscis also serves as the developmental site for the infective metacyclic stages of several species of pathogenic livestock trypanosomes that are inoculated into the host with fly saliva. To understand the physical environment in which these trypanosomes develop, we have re-examined the microarchitecture of the tsetse proboscis. RESULTS: We examined proboscises from male and female flies of Glossina pallidipes using light microscopy and scanning electron microscopy (SEM). Each proboscis was removed from the fly head and either examined intact or dissected into the three constituent components: Labrum, labium and hypopharynx. Our light and SEM images reaffirm earlier observations that the tsetse proboscis is a formidably armed weapon, well-adapted for piercing skin, and provide comparative data for G. pallidipes. In addition, the images reveal that the hypopharynx, the narrow tube that delivers saliva to the wound site, ends in a remarkably ornate and complex structure with around ten finger-like projections, each adorned with sucker-like protrusions, contradicting previous descriptions that show a simple, bevelled end like a hypodermic needle. The function of the finger-like projections is speculative; they appear to be flexible and may serve to protect the hypopharynx from influx of blood or microorganisms, or control the flow of saliva. Proboscises were examined after colonisation by Trypanosoma congolense savannah. Consistent with the idea that colonisation commences in the region nearest the foregut, the highest densities of trypanosomes were found in the region of the labrum proximal to the bulb, although high densities were also found in other regions of the labrum. Trypanosomes were visible through the thin wall of the hypopharynx by both light microscopy and SEM. CONCLUSIONS: We highlight the remarkable architecture of the tsetse proboscis, in particular the intricate structure of the distal end of the hypopharynx. Further work is needed to elucidate the function of this intriguing structure.

Genetic and morphometric evidence for population isolation of Glossina palpalis gambiensis (Diptera: Glossinidae) on the Loos islands, Guinea.

Allele frequencies at four microsatellite loci, and morphometric features based on 11 wing landmarks, were compared among three populations of Glossina palpalis gambiensis (Diptera: Glossinidae) in Guinea. One population originated from the Loos islands separated from the capital Conakry by 5 km of sea, and the two others originated from the continental mangrove area close to Dubreka, these two groups being separated by approximately 30 km. Microsatellites and wing geometry data both converged to the idea of a separation of the Loos island population from those of the mangrove area. Although occasional contacts cannot be excluded, our results support the hypothesis of the Loos population of tsetse flies being a completely isolated population. This situation will favor a sequenced intervention against human African trypanosomosis and the possibility of an elimination of tsetse from this island.

Multiple procyclin isoforms are expressed differentially during the development of insect forms of Trypanosoma brucei.

Transmission of Trypanosoma brucei by the tsetse fly entails several rounds of differentiation as the parasite migrates through the digestive tract to the salivary glands of its vector. Differentiation of the bloodstream to the procyclic form in the fly midgut is accompanied by the synthesis of a new coat consisting of EP and GPEET procyclins. There are three closely related EP isoforms, two of which (EP1 and EP3) contain N-glycans. To identify the individual EP isoforms that are expressed early during synchronous differentiation in vitro, we exploited the selective extraction of GPI-anchored proteins and mass spectrometry. Unexpectedly, we found that GPEET and all isoforms of EP were coexpressed for a few hours at the onset of differentiation. At this time, the majority of EP1 and EP3 molecules were already glycosylated. Within 24 hours, GPEET became the major surface component, to be replaced in turn by glycosylated forms of EP, principally EP1, at a later phase of development. Transient transfection experiments using reporter genes revealed that each procyclin 3' untranslated region contributes to differential expression as the procyclic form develops. We postulate that programmed expression of other procyclin species will accompany further rounds of differentiation, enabling the parasite to progress through the fly.

Peptidomics of Neuropeptidergic Tissues of the Tsetse Fly Glossina morsitans morsitans.

Neuropeptides and peptide hormones are essential signaling molecules that regulate nearly all physiological processes. The recent release of the tsetse fly genome allowed the construction of a detailed in silico neuropeptide database (International Glossina Genome Consortium, Science 344, 380-386 (2014)), as well as an in-depth mass spectrometric analysis of the most important neuropeptidergic tissues of this medically and economically important insect species. Mass spectrometric confirmation of predicted peptides is a vital step in the functional characterization of neuropeptides, as in vivo peptides can be modified, cleaved, or even mispredicted. Using a nanoscale reversed phase liquid chromatography coupled to a Q Exactive Orbitrap mass spectrometer, we detected 51 putative bioactive neuropeptides encoded by 19 precursors: adipokinetic hormone (AKH) I and II, allatostatin A and B, capability/pyrokinin (capa/PK), corazonin, calcitonin-like diuretic hormone (CT/DH), FMRFamide, hugin, leucokinin, myosuppressin, natalisin, neuropeptide-like precursor (NPLP) 1, orcokinin, pigment dispersing factor (PDF), RYamide, SIFamide, short neuropeptide F (sNPF) and tachykinin. In addition, propeptides, truncated and spacer peptides derived from seven additional precursors were found, and include the precursors of allatostatin C, crustacean cardioactive peptide, corticotropin releasing factor-like diuretic hormone (CRF/DH), ecdysis triggering hormone (ETH), ion transport peptide (ITP), neuropeptide F, and proctolin, respectively. The majority of the identified neuropeptides are present in the central nervous system, with only a limited number of peptides in the corpora cardiaca-corpora allata and midgut. Owing to the large number of identified peptides, this study can be used as a reference for comparative studies in other insects. Graphical Abstract ᅟ.

Artificial warthog burrows used to sample adult and immature tsetse (Glossina spp) in the Zambezi Valley of Zimbabwe.

BACKGROUND: The biology of adult tsetse (Glossina spp), vectors of trypanosomiasis in Africa, has been extensively studied - but little is known about larviposition in the field. METHODOLOGY/PRINCIPAL FINDINGS: In September-November 1998, in the hot-dry season in Zimbabwe's Zambezi Valley, we used artificial warthog burrows to capture adult females as they deposited larvae. Females were subjected to ovarian dissection and were defined as perinatal flies, assumed to have entered burrows to larviposit, if oocyte sizes indicated >95% pregnancy completion. Perinatal flies were defined as full-term pregnant if there was a late third instar larva in utero, or postpartum if the uterus was empty. All other females were defined as pre-full-term pregnant (pre-FT). Of 845 G. m. morsitans captured, 91% (765) were female and 295/724 (41%) of females dissected were perinatal flies. By contrast, of 2805 G. pallidipes captured only 71% (2003) were female and only 33% (596/1825) of females were perinatal. Among all perinatal females 67% (596/891) were G. pallidipes. Conversely, in burrows not fitted with traps - such that flies were free to come and go - 1834 (59%) of pupae deposited were G. m. morsitans and only 1297 (41%) were G. pallidipes. Thus, while more full-term pregnant G. pallidipes enter burrows, greater proportions of G. m. morsitans larviposit in them, reflecting a greater discrimination among G. pallidipes in choosing larviposition sites. Catches of males and pre-FT females increased strongly with temperatures above 32°C, indicating that these flies used burrows as refuges from high ambient temperatures. Conversely, catches of perinatal females changed little with maximum temperature but declined from late September through November: females may anticipate that burrows will be inundated during the forthcoming wet season. Ovarian age distributions of perinatal and pre-FT females were similar, consistent with all ages of females larvipositing in burrows with similar probability. CONCLUSIONS/SIGNIFICANCE: Artificial warthog burrows provide a novel method for collecting tsetse pupae, studying tsetse behaviour at larviposition, assessing the physiological status of female tsetse and their larvae, and of improving understanding of the physiological dynamics of terminal pregnancy, and population dynamics generally, with a view to improving methods of trypanosomiasis control.

Left-right asymmetry of fly wings and the evolution of body axes.

The body plan of Drosophila, and presumably that of other insects, develops under the control of anterio-posterior and dorsal ventral axes, but no evidence for a left-right axis has yet been found. We used geometric morphometrics to study the wings in three species of flies: Drosophila melanogaster, Musca domestica and Glossina palpalis gambiensis. In all three species, we found that both size and shape showed subtle, but statistically significant directional asymmetry. For size, these asymmetries were somewhat inconsistent within and between species, but for shape, highly significant directional asymmetry was found in all samples examined. These systematic left-right differences imply the existence of a left-right axis that conveys distinct positional identities to the wing imaginal discs on either body side. Hence, the wing discs of Drosophila may be a new model to study the developmental genetics of left-right asymmetry. The asymmetries of shape were similar among species, suggesting that directional asymmetry has been evolutionarily conserved since the three lineages diverged. We discuss the implications of this evolutionary conservatism in conjunction with results from earlier studies that showed a lack of genetic variation for directional asymmetry in Drosophila.

A preliminary investigation of the effect of age, sex and time of collection on the feeding patterns of Glossina morsitans morsitans Westw. in Zambia.

Glossina morsitans morsitans were collected during the dry season of 1975 from two areas in the Luangwa Valley (Zambia) and from one of the areas in the wet season of 1976. In all, 1,190 flies were analysed for sex, wing fray category and source of bloodmeal. Differences in the feeding patterns in the morning and afternoon collections reflected host behaviour. Warthog consistently emerged as a major host but there appeared to be some local variation resulting from seasonal and diurnal availability of hosts.