Mass irradiation of adult Aedes mosquitoes using a coolable 3D printed canister.

In the last decade, the use of the sterile insect technique (SIT) to suppress mosquito vectors have rapidly expanded in many countries facing the complexities of scaling up production and procedures to sustain large-scale operational programs. While many solutions have been proposed to improve mass production, sex separation and field release procedures, relatively little attention has been devoted to effective mass sterilization of mosquitoes. Since irradiation of pupae en masse has proven difficult to standardise with several variables affecting dose response uniformity, the manipulation of adult mosquitoes appears to be the most promising method to achieve effective and reliable sterilization of large quantities of mosquitoes. A 3D-printed phase change material based coolable canister was developed which can compact, immobilize and hold around 100,000 adult mosquitoes during mass radio sterilization procedures. The mass irradiation and compaction treatments affected the survival and the flight ability of Aedes albopictus and Aedes aegypti adult males but the use of the proposed irradiation canister under chilled conditions (6.7-11.3 °C) significantly improved their quality and performance. The use of this cooled canister will facilitate adult mass irradiation procedures in self-contained irradiators in operational mosquito SIT programmes.

Field performance of sterile male mosquitoes released from an uncrewed aerial vehicle.

Genetic control methods of mosquito vectors of malaria, dengue, yellow fever, and Zika are becoming increasingly popular due to the limitations of other techniques such as the use of insecticides. The sterile insect technique is an effective genetic control method to manage insect populations. However, it is crucial to release sterile mosquitoes by air to ensure homogeneous coverage, especially in large areas. Here, we report a fully automated adult mosquito release system operated from an uncrewed aerial vehicle or drone. Our system, developed and tested in Brazil, enabled a homogeneous dispersal of sterile male Aedes aegypti while maintaining their quality, leading to a homogeneous sterile-to-wild male ratio due to their aggregation in the same sites. Our results indicate that the released sterile males were able to compete with the wild males in mating with the wild females; thus, the sterile males were able to induce sterility in the native female population. The use of drones to implement the sterile insect technique will lead to improvements in areal coverage and savings in operational costs due to the requirement of fewer release sites and field staff.

Genetic comparison of Glossina tachinoides populations in three river basins of the Upper West Region of Ghana and implications for tsetse control.

Tsetse flies are the cyclical vectors of African animal trypanosomosis (AAT) and human African trypanosomosis (HAT). In March 2010, the Government of Ghana initiated a large scale integrated tsetse eradication campaign in the Upper West Region (UWR) (≈18,000 km(2)) under the umbrella of the Pan-African Tsetse and Trypanosomosis Eradication Campaign (PATTEC). We investigated the structuring of Glossina tachinoides populations within and between the three main river basins of the target area in the UWR. Out of a total sample of 884 flies, a sub-sample of 266 was genotyped at nine microsatellite loci. The significance of the different hierarchical levels was tested using Yang's parameters estimated with Weir and Cockerham's method. A significant effect of traps within groups (pooling traps no more than 3 km distant from each other), of groups within river basins and of river basins within the whole target area was observed. Isolation by distance between traps was highly significant. A local density of 0.48-0.61 flies/m(2) was estimated and a dispersal distance that approximated 11 m per generation [CI 9, 17]. No significant sex-biased dispersal was detected. Dispersal distances of G. tachinoides in the UWR were relatively low, possibly as a result of the fragmentation of the habitat and the seasonality of the Kulpawn and Sissili rivers. Moreover, very high fly population densities were observed in the sample sites, which potentially reduces dispersal at constant habitat saturation, because the probability that migrants can established is reduced (density dependent dispersal). However, the observed spatial dispersal was deemed sufficient for a G. tachinoides-cleared area to be reinvaded from neighboring populations in adjacent river basins. These data corroborate results from other population genetics studies in West Africa, which indicate that G. tachinoides populations from different river basins cannot be considered isolated.

Mating compatibility between Bactrocera invadens and Bactrocera dorsalis (Diptera: Tephritidae).

The invasive fruit fly, Bactrocera invadens Drew, Tsuruta & White, is a highly polyphagous fruit pest that occurs predominantly in Africa yet has its origins in the Indian subcontinent. It is extremely morphologically and genetically similar to the Oriental fruit fly, Bactrocera dorsalis (Hendel); as such the specific relationship between these two species is unresolved. We assessed prezygotic compatibility between B. dorsalis and B. invadens using standardized field cage mating tests, which have proven effectiveness in tephritid cryptic species studies. These tests were followed by an assessment of postzygotic compatibility by examining egg viability, larval and pupal survival, and sex ratios of offspring produced from parental and subsequent F1 crosses to examine for hybrid breakdown as predicted under a two-species hypothesis. B. dorsalis was sourced from two countries (Pakistan and China), and each population was compared with B. invadens from its type locality of Kenya. B. invadens mated randomly with B. dorsalis from both localities, and there were generally high levels of hybrid viability and survival resulting from parental and F1 crosses. Furthermore, all but one hybrid cross resulted in equal sex ratios, with the single deviation in favor of males and contrary to expectations under Haldane's rule. These data support the hypothesis that B. dorsalis and B. invadens represent the same biological species, an outcome that poses significant implications for pest management and international trade for sub-Saharan Africa.

Mating compatibility among four pest members of the Bactrocera dorsalis fruit fly species complex (Diptera: Tephritidae).

Bactrocera dorsalis (Hendel), Bactrocera papayae Drew & Hancock, Bactrocera philippinensis Drew & Hancock, and Bactrocera carambolae Drew & Hancock are pest members within the B. dorsalis species complex of tropical fruit flies. The species status of these taxa is unclear and this confounds quarantine, pest management, and general research. Mating studies carried out under uniform experimental conditions are required as part of resolving their species limits. These four taxa were collected from the wild and established as laboratory cultures for which we subsequently determined levels of prezygotic compatibility, assessed by field cage mating trials for all pair-wise combinations. We demonstrate random mating among all pair-wise combinations involving B. dorsalis, B. papayae, and B. philippinensis. B. carambolae was relatively incompatible with each of these species as evidenced by nonrandom mating for all crosses. Reasons for incompatibility involving B. carambolae remain unclear; however, we observed differences in the location of couples in the field cage for some comparisons. Alongside other factors such as pheromone composition or other courtship signals, this may lead to reduced interspecific mating compatibility with B. carambolae. These data add to evidence that B. dorsalis, B. papayae, and B. philippinensis represent the same biological species, while B. carambolae remains sufficiently different to maintain its current taxonomic identity. This poses significant implications for this group's systematics, impacting on pest management, and international trade.

Anopheles arabiensis sperm production after genetic manipulation, dieldrin treatment, and irradiation.

The use of the sterile insect technique relies on the release of sterilized mass-reared male insects which, before field releases, endure several unnatural treatments. In the case of Anopheles arabiensis (Patton) sterile insect technique program in Sudan, the genetic background of the original strain was first changed to create a genetic sexing strain that is based on a dieldrin-resistant mutation. Secondly, the eggs of the genetic sexing strain require treatment with dieldrin to allow complete elimination of female L1 larvae to enable the release of males only. Finally, male mosquitoes receive an irradiation dose of 70 Gy as pupae for sterilization. The effects of these treatments on sperm production were tested separately and in combination. Irradiation alone significantly decreased the initial sperm number and prevented new sperm production. However, the dieldrin treatment, aimed at eliminating females, appears to have an unexpected radioprotectant effect.

Different blood and sugar feeding regimes affect the productivity of Anopheles arabiensis colonies (Diptera: Culicidae).

The success of the sterile insect technique for the management of mosquito populations depends on the release of large numbers of competitive sterile male insects. Sustainable mosquito production can only be obtained when proper mass-rearing equipment and adequate methods are available, including those to feed blood to the female mosquitoes. The blood feeding apparatus Hemotek consists of a small aluminum plate to which a collagen membrane is fixed and filled with blood kept warm by an electric heating element. A larger aluminum plate was developed to feed a larger number of female mosquitoes with blood that is kept at a constant temperature. The effect of different blood feeding regimes (feeding frequency and time the blood is kept in the Hemotek) and sugar deprivation before blood feeding on egg production of female Anopheles arabiensis Patton was tested. Egg production was higher when blood was offered to the mosquitoes every day as compared with every 2 or 4 d. Sugar deprivation for 7 h before blood feeding enhanced egg production by 50% compared with female mosquitoes that had continuous access to sugar. Neither male nor female survival was impaired. Finally, we showed that the same blood could be kept warm and used over several hours to feed mosquitoes in multiple cages without any impact on egg production or hatch rate. Being able to use the same blood over extended periods would save considerable time, handling, and funds.

Prevalence and genetic variation of salivary gland hypertrophy virus in wild populations of the tsetse fly Glossina pallidipes from southern and eastern Africa.

The Glossina pallidipes salivary gland hypertrophy virus (GpSGHV) is a rod-shaped, non-occluded double-stranded DNA virus that causes salivary gland hypertrophy (SGH) and reduced fecundity in the tsetse fly G. pallidipes. High GpSGHV prevalence (up to 80%) makes it impossible to mass-rear G. pallidipes colonies for the sterile insect technique (SIT). To evaluate the feasibility of molecular-based GpSGHV management strategies, we investigated the prevalence and genetic diversity of GpSGHV in wild populations of G. pallidipes collected from ten geographical locations in eastern and southern Africa. Virus diversity was examined using a total sequence of 1497 nucleotides (≈ 1% of the GpSGHV genome) from five putative conserved ORFs, p74, pif1, pif2, pif3 and dnapol. Overall, 34.08% of the analyzed flies (n=1972) tested positive by nested PCR. GpSGHV prevalence varied from 2% to 100% from one location to another but phylogenetic and gene genealogy analyses using concatenated sequences of the five putative ORFs revealed low virus diversity. Although no correlation of the virus diversity to geographical locations was detected, the GpSGHV haplotypes could be assigned to one of two distinct clades. The reference (Tororo) haplotype was the most widely distributed, and was shared by 47 individuals in seven of the 11 locations. The Ethiopian haplotypes were restricted to one clade, and showed the highest divergence (with 14-16 single nucleotide mutation steps) from the reference haplotype. The current study suggests that the proposed molecular-based virus management strategies have a good prospect of working throughout eastern and southern Africa due to the low diversity of the GpSGHV strains.

Quality of mass-reared codling moth (Lepidoptera: Tortricidae) after long-distance transportation: 1. Logistics of shipping procedures and quality parameters as measured in the laboratory.

The sterile insect technique (SIT) is a proven effective control tactic against lepidopteran pests when applied in an areawide integrated pest management program. The construction of insect mass-rearing facilities requires considerable investment and moth control strategies that include the use of sterile insects could be made more cost-effective through the importation of sterile moths produced in other production centers. For codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae), this is an attractive option because mating studies have confirmed the absence of mating barriers between codling moth populations from geographically different areas. To assess the feasibility of long-distance transportation of codling moths, pupae and adult moths were transported in 2004 from Canada to South Africa in four shipments by using normal commercial transport routes. The total transport time remained below 67 h in three of the consignments, but it was 89 h in the fourth consignment. Temperature in the shipping boxes was fairly constant and remained between -0.61 and 0.16 degrees C for 76.8-85.7% of the time. The data presented indicate that transporting codling moths as adults and pupae from Canada to South Africa had little effect on moth emergence, longevity, and ability to mate, as assessed in the laboratory. These results provide support to the suggestion that the STT for codling moth in pome fruit production areas might be evaluated and implemented by the importation of irradiated moths from rearing facilities in a different country or hemisphere.

The prevalence of African animal trypanosomoses and tsetse presence in Western Senegal.

In 2005, the Government of Senegal initiated a tsetse eradication campaign in the Niayes and La Petite Côte aiming at the removal of African Animal Trypanosomosis (AAT), which is one of the main constraints to the development of more effective cattle production systems. The target area has particular meteorological and ecological characteristics that provide great potential for animal production, but it is unfortunately still infested by the riverine tsetse species Glossina palpalis gambiensis Vanderplank (Diptera: Glossinidae). The tsetse project in Senegal has adopted an area-wide integrated pest management (AW-IPM) approach that targets the entire tsetse population within a delimited area. During the first phase of the programme, a feasibility study was conducted that included the collection of entomological, veterinary, population genetics, environmental and socioeconomic baseline data. This paper presents the parasitological and serological prevalence data of AAT in cattle residing inside and outside the tsetse-infested areas of the target zone prior to the control effort. At the herd level, a mean parasitological prevalence of 2.4% was observed, whereas a serological prevalence of 28.7%, 4.4%, and 0.3% was obtained for Trypanosoma vivax, T. congolense and T. brucei brucei, respectively. The observed infection risk was 3 times higher for T. congolense and T. vivax in the tsetse-infested than in the assumed tsetse-free areas. Moreover, AAT prevalence decreased significantly with distance from the nearest tsetse captured which indicated that cyclical transmission of the parasites by tsetse was predominant over mechanical transmission by numerous other biting flies present. The importance of these results for the development of a control strategy for the planned AW-IPM campaign is discussed.

Enabling technologies to improve area-wide integrated pest management programmes for the control of screwworms.

The economic devastation caused in the past by the New World screwworm fly Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae) to the livestock industry in the U.S.A., Mexico and the rest of Central America was staggering. The eradication of this major livestock pest from North and Central America using the sterile insect technique (SIT) as part of an area-wide integrated pest management (AW-IPM) programme was a phenomenal technical and managerial accomplishment with enormous economic implications. The area is maintained screwworm-free by the weekly release of 40 million sterile flies in the Darien Gap in Panama, which prevents migration from screwworm-infested areas in Columbia. However, the species is still a major pest in many areas of the Caribbean and South America and there is considerable interest in extending the eradication programme to these countries. Understanding New World screwworm fly populations in the Caribbean and South America, which represent a continuous threat to the screwworm-free areas of Central America and the U.S.A., is a prerequisite to any future eradication campaigns. The Old World screwworm fly Chrysomya bezziana Villeneuve (Diptera: Calliphoridae) has a very wide distribution ranging from Southern Africa to Papua New Guinea and, although its economic importance is assumed to be less than that of its New World counterpart, it is a serious pest in extensive livestock production and a constant threat to pest-free areas such as Australia. In the 1980s repeated introductions and an expansion of Old World screwworm populations were reported in the Middle East; in the 1990s it invaded Iraq and since late 2007 it has been reported in Yemen, where a severe outbreak of myiasis occurred in 2008. Small-scale field trials have shown the potential of integrating the SIT in the control of this pest and various international organizations are considering using the release of sterile insects as part of an AW-IPM approach on a much wider scale. Wohlfahrtia magnifica (Schiner) (Diptera: Sarcophagidae) is a screwworm of temperate regions, which, although of limited agricultural importance, has invaded several new locations in the past few years. This special issue reports on the results of a 6-year project funded by the Joint Food and Agriculture Organization of the United Nations/International Atomic Energy Agency (FAO/IAEA) Programme of Nuclear Techniques in Food and Agriculture entitled 'Enabling Technologies for the Expansion of the SIT for Old and New World Screwworm'. A major goal of the project was to better understand population genetic variation in screwworms as an aid to the identification of isolated populations. The project also addressed issues related to genetic sexing, cuticular hydrocarbons, population dynamics, genetic transformation and chromosome analysis.

Mitochondrial diversity analysis of Glossina palpalis gambiensis from Mali and Senegal.

West African riverine tsetse populations of Glossina palpalis gambiensis Vanderplank (Diptera: Glossinidae) were investigated for gene flow, inferred from mitochondrial diversity in samples of 69 flies from Senegal and 303 flies from three river drainages in Mali. Four polymorphic mitochondrial loci were scored. Mean haplotype diversities were 0.30 in Mali, 0 in Senegal and 0.18 over both Mali and Senegal. These diversities estimate the probabilities that two randomly chosen tsetse have different haplotypes. Substantial rates of gene flow were detected among flies sampled along tributaries belonging to the river basins of the Senegal, Niger, and Bani in Mali. There was virtually no gene flow between tsetse in Senegal and Mali. No seasonal effects on gene flow were detected. The implications of these preliminary findings for the implementation of area-wide integrated pest management (AW-IPM) programmes against riverine tsetse in West Africa are discussed.

Long-term sampling of gamma sterilised male Glossina austeni (Diptera: Glossinidae) with sticky panels on Unguja Island (Zanzibar).

Daily catches of gamma sterilised male Glossina austeni Newstead with experimental sticky panels were analysed from March 1996 to July 1997. The flies were released weekly by light aircraft over primary and secondary forest ecosystems of Unguja Island, Zanzibar. In the primary forest, the cross-shaped royal blue XT panel (two interlocking panels of each 70 x 60 cm) trapped significantly more flies than the royal blue-white leg panel (panel with a body of 65 x 30 cm and two legs of each 15 x 15 cm) in all months, except in July 1996 and July 1997. In the same habitat, the cross-shaped royal blue-white leg panel (two interlocking leg panels) trapped from 1.7 (not significant) to 3.0 times (highly significant) as many flies as the standard leg panel depending on the season. Significantly more flies were trapped with the cross-shaped XT panel than with the leg panel in the secondary forest compared to the primary forest. Catches of the cross-shaped XT panel on each of a series of days, correlated well with those of the leg panel on the same day, except for panels deployed in the primary forest during the hot-dry and the beginning of the cold-dry season. Catches of the cross-shaped leg panel correlated less well with the catches of the standard leg panel in the primary forest. The data presented indicate that the behavioural responses in time and space of sterile male G. austeni are influenced by the type of trapping device used.

Principles of area-wide integrated tsetse fly control using the sterile insect technique.

The tsetse fly and the disease trypanosomosis it transmits, is one of the most severe medical and veterinary problems in Africa, infecting around 50,000 people every year and preventing the development of sustainable and productive agricultural systems. The most efficient way to contain the disease is by the management of entire populations of the vector (area-wide approach) using a combination of several control methods in an integrated pest management campaign. A very powerful method for integration in these programmes as a final eradication component is the sterile insect technique (SIT). The technique relies on the rearing of the target insect in large numbers in specialised production centres, the sterilisation with ionising radiation of one of the sexes and the sustained sequential release of the sterilised insects over the target area. Contrary to conventional control methods, the sterile insect technique becomes more efficient with decreasing density of the target population. Hence, the technique fits well within the concept of integrated pest management, as its complementary use in a phased approach with other suppression techniques, results in maximum efficiency. The five main prerequisites required to use the sterile insect technique against a target insect have been achieved for tsetse flies: 1. adequate information on the ecology of the insect is available, 2. the mass-production of the target insect is economically feasible, 3. efficient suppression techniques are available to reduce the population density of the target insect, 4. sufficient sterile males are available for release and their competitiveness is optimal, 5. the gamma treated sperm is competitive with the wild sperm. This paper gives a detailed review of the five implementation phases of the sterile insect technique based area-wide integrated pest management campaign against Glossina austeni on the island of Unguja, Zanzibar. The potential future prospects and requirements of using the sterile insect technique against tsetse populations are likewise presented.

Glossina austeni (Diptera: Glossinidae) eradicated on the island of Unguja, Zanzibar, using the sterile insect technique.

An area-wide integrated tsetse eradication project was initiated in Zanzibar in 1994 by the International Atomic Energy Agency and the governments of Tanzania and Zanzibar, to eradicate Glossina austeni Newstead from Unguja Island (Zanzibar) using the sterile insect technique. Suppression of the tsetse population on Unguja was initiated in 1988 by applying residual pyrethroids as a pour-on formulation to livestock and by the deployment of insecticide impregnated screens in some of the forested areas. This was followed by sequential releases of gamma-sterilized male flies by light aircraft. The flies, packaged in carton release containers, were dispersed twice a week along specific flight lines separated by a distance of 1-2 km. More than 8.5 million sterile male flies were released by air from August 1994 to December 1997. A sterile to indigenous male ratio of >50:1 was obtained in mid-1995 and it increased to >100:1 by the end of 1995. As a consequence the proportion of sampled young females (1-2 ovulations), with an egg in utero in embryonic arrest or an uterus empty as a result of expulsion of a dead embryo, increased from <25% in the 1st quarter to >70% in the last quarter of 1995. In addition, the age structure of the female population became significantly distorted in favor of old flies (> or = 4 ovulations) by the end of 1995. The apparent density of the indigenous fly population declined rapidly in the last quarter of 1995, followed by a population crash in the beginning of 1996. The last trapped indigenous male and female flies were found in weeks 32 and 36, 1996, respectively. Time for 6 fly generations elapsed between the last catch of an indigenous fly and the end of the sterile male releases in December 1997.

Field responses of Glossina austeni to sticky panels on Unguja Island, Zanzibar.

Two designs of cross-shaped sticky panels (XT and XLP) were compared with the royal blue-white legpanel (LPBuWh) in the Jozani forest on Unguja Island as trapping devices for male Glossina austeni. Single coloured royal blue (XTBu) and bi-coloured royal blue-white XT (XTBuWh) caught more than twice as many male G. austeni as the LPBuWh, whereas single coloured black XT trapped significantly fewer flies (10%) than the control LPBuWh. XT's in various horizontal and diagonal blue-white configurations likewise trapped more flies than the LPBuWh, except a horizontally striped blue-white XT which trapped fivefold fewer flies than the LPBuWh. Cross-shaped LP in the blue-white (XLPBuWh) and black-white (XLPB1Wh) combination scored significantly better than the control LPBuWh. Similar fly numbers were trapped with XTBuWh and XLPBuWh. Long-term trapping data indicated that the XTBu, XTBuWh and XLPBuWh were three- to fourfold more effective in trapping female flies than the LPBuWh. The landing bias on bi-coloured panels was low in the blue-white but more pronounced in the blue-black and white-black combinations and was affected by the type of sticky panel used. A high proportion (49%) of the flies alighted on the bottom corners of the XTBu panel, but landing positions were more scattered if white was added. Increasing the width of the XTBu from 70 to 120 cm improved the catch by a factor of two as compared with standard sized XTBu. The effect of doubling the height of the XT on total fly catch was negligible. At present, it is the XTBu which can be recommended as the best trapping device for male and female G. austeni.

Comparative gamma-radiation sensitivity of Glossina tachinoides Westv., Glossina fuscipes fuscipes Newst. and Glossina brevipalpis Newst. (Diptera, Glossinidae).

The effect of gamma-radiation doses ranging between 10 and 180 Gy on 4-6-day-old adult males of Glossina tachinoides, Glossina fuscipes fuscipes and Glossina brevipalpis was studied. Fecundity of their mates was reduced by 95% following exposure to 120, 80-100 and 50 Gy of adult male G. tachinoides, G. f. fuscipes and G. brevipalpis respectively. Insemination ability of the males and sperm motility were not adversely affected by the radiation treatment. The higher proportion of dominant lethals in the sperm of the three species with increasing radiation doses was reflected in the reproductive status of the female mates, i.e. an increasing percentage of females showing imbalances between intrauterine content and ovarian development (females with an empty uterus due to expulsion of a dead embryo after embryonic arrest or a degenerating egg in utero) and an acceleration in follicle development associated with successive unsuccessful cycles. In the F1 progeny of all treated groups, no significant bias of the sex ratio was found. The average life span of G. tachinoides and G. f. fuscipes males treated with doses of > or = 80 Gy and of G. brevipalpis males treated with doses >140 Gy was significantly reduced as compared with untreated controls. Male G. brevipalpis treated with doses ranging between 10 and 40 Gy, however, showed a significant radiation induced increase in average life span.

Analysis of the mating scar pattern of Glossina palpalis palpalis (Rob.-Desv.) and Glossina fuscipes fuscipes Newstead (Diptera: Glossinidae).

An analysis was made of the mating scar pattern of female Glossina palpalis palpalis Robineau-Desvoidy and Glossina fuscipes fuscipes Newstead. Measurements on fifty permanent preparations of the mating scars of females reared in the laboratory revealed significant differences in the length, width and in the distance between the centers of the mating scars of the two species. Plotting the distance between the centers of the two mating scars against the ratio width/length resulted in a 93% separation of the two species. It is proposed that this technique could be used during field surveys to expose possible cross-breeding in nature or as a tool in the entomological evaluation of a tsetse eradication campaign where one species is released in the habitat of the other.

The effect of intersubspecific hybridization and gamma radiation on the reproductive biology of Glossina palpalis palpalis (Robineau-Desvoidy) and Glossina palpalis gambiensis Vanderplank.

The closely related tsetse fly subspecies Glossina palpalis palpalis (Nigeria origin) and Glossina palpalis gambiensis (Burkina Faso origin) hybridize readily in the laboratory. Hybridized G.p.palpalis females produced less offspring than the parental intrasubspecific crosses. Adult emergence was below 70% with at least 78% being females. Most female hybrids were fertile whereas most of the male hybrids were sterile when backcrossed to the G.p.palpalis parental line. All F1 males were capable of transferring a spermatophore but their mates rarely had sperm-impregnated spermathecae. Their testes rarely contained mature sperm; moreover, sperm, when present had low or no motility. During laboratory cage tests with virgin females of both subspecies and either sexually mature male G.p.palpalis or G.p.gambiensis, there was no indication for selective mating. The same was true when gamma irradiated males (120 Gy treatment in air) were used. In the latter case complete sterility was induced causing embryonic arrest in all inseminated female mates. Consequently, in ratio tests with untreated virgin G.p.palpalis females, untreated G.p.palpalis males and an increasing number of irradiated G.p.gambiensis males, there was a gradual decrease in production of viable offspring. The results of the present study are discussed with a view of using a combined hybridization and induced sterility in distinct geographical zones where the two subspecies are present.