Mating compatibility and competitiveness of transgenic and wild type Aedes aegypti (L.) under contained semi-field conditions

We conducted the world??s first experiments under semi-field conditions (ACL-2 field house) to assess the mating competitiveness of genetically sterile RIDL male mosquitoes (513A strain). The field house is a state-of-the-art, fully-contained trial facility, simulating the living space for a household of 2?C4 people in Peninsular Malaysia. Ten genetically sterile RIDL male A. aegypti mosquitoes competed with ten wild type males inside this field house to mate with ten wild type females. Hatched larvae from mated females were screened under a fluorescent microscope for genetic markers to determine if they were fathered by RIDL male or wild type male, and all results were cross-checked by PCR. Two such experiments were conducted, each repeated sufficient number of times. All strains were on a Malaysian lab strain background for the first experiment, while the RIDL males alone were on a recently-colonised Mexican strain background for the second experiment. A total of 52 % of the matings were with RIDL males in the first experiment, while 45 % of the matings were with RIDL (Mexican) males in the second experiment. Statistically, this is not significantly different from 50 % of the matings expected to take place with RIDL males if the latter were as competitive as that of the wild type males. This shows that A. aegypti RIDL-513A has excellent mating competitiveness under semi-field conditions, verifying earlier trends obtained in small lab cages. We also observed high mating compatibility between recently-colonised Mexican RIDL males and lab-reared Malaysian wild type females.     

Male lures and male mating competitiveness in a genetic sexing strain of the melon fly (Diptera: Tephritidae)

The Sterile Insect Technique (SIT) has been used to suppress or eradicate populations of the melon fly, Zeugodacus cucurbitae (Coquillett), a pest of various fruits and vegetables throughout Asia, Africa, and South Pacific Islands. The success of the SIT depends largely on the ability of released, sterile males to compete successfully with wild males for copulations with wild females. Unfortunately, both the sterilization process (via gamma irradiation) and artificial selection under mass-rearing conditions generally reduce the biological competence, including the mating competitiveness, of the released males. However, previous data on Z. cucurbitae show that irradiation has minimal adverse effects and prolonged mass-rearing may or may not result in decreased mating ability. The present study first confirms that neither irradiation nor mass-rearing had deleterious effects on male mating ability of males from a pupal-color, genetic sexing strain (termed T1). Then, the influence of male lures (cue-lure and its natural analogue raspberry ketone) on the mating success of sterile T1 males was measured as a potential pre-release procedure to improve the effectiveness of SIT. Feeding on cue-lure enhanced male mating success, but the effect was short-lived (1 d but not 3 d post-feeding). Feeding on raspberry ketone boosted mating success over a longer time, i.e., at least 5 d but not 10 d post-feeding. However, mixing raspberry ketone into the adult diet had no detectable effect on male mating performance. The implications of these results for SIT against Z. cucurbitae are discussed.     

Repellent efficacy of wood vinegar against Culex pipiens pallens and Aedes togoi (Diptera: Culicidae) under laboratory and semi-field conditions

The repellent efficacy of wood vinegar was assessed against mosquitoes under laboratory conditions at 1, 5, 10, 20, 40, 60 and 80% concentrations. The study evaluated whether wood vinegar is able to repel Culex pipiens pallens Coquillet and Aedes togoi (Theobald) from the human body and if so at what concentrations. The tests were conducted using the arm-in-cage method in 80 × 40 × 40 cm screened mosquito cages. The data were analyzed and compared with those of N,N-Diethyl-3-methylbenzamide (deet) at 10.3% concentration. The results showed that wood vinegar provided mosquito repellence of varying degree depending on the concentration used. The observed repellence averaged from as low as 39.6% at 5.0% concentration to as high as 100% at 80% concentration against Ae. togoi. Repellence against Cx. pipiens pallens was high being 90.3% at 20% concentration, 92.2% at 40% concentration, 93.9% at 60% concentration and 100% at 80% concentration. The duration of protection time tests showed that the 40% and 60% concentrations of the wood vinegar give protection from landing of Ae. togoi for a period of up to 7 h, though the lower concentration gave lower protection after the first five hours. The results indicated that wood vinegar has mosquito repellent characteristics that tend to vary with the concentration used and the species of mosquitoes. Wood vinegar in this case was very effective in repelling Cx. pipiens pallens, even at lower concentrations while higher concentrations were required to repel Ae. togoi.     

A secure semi-field system for the study of Aedes aegypti

Background

New contained semi-field cages are being developed and used to test novel vector control strategies of dengue and malaria vectors. We herein describe a new Quarantine Insectary Level-2 (QIC-2) laboratory and field cages (James Cook University Mosquito Research Facility Semi-Field System; MRF SFS) that are being used to measure the impact of the endosymbiont Wolbachia pipientis on populations of Aedes aegypti in Cairns Australia.

Methodology/Principal Findings

The MRF consists of a single QIC-2 laboratory/insectary that connects through a central corridor to two identical QIC-2 semi-field cages. The semi-field cages are constructed of two layers of 0.25 mm stainless steel wire mesh to prevent escape of mosquitoes and ingress of other insects. The cages are covered by an aluminum security mesh to prevent penetration of the cages by branches and other missiles in the advent of a tropical cyclone. Parts of the cage are protected from UV light and rainfall by 90% shade cloth and a vinyl cover. A wooden structure simulating the understory of a Queenslander-style house is also situated at one end of each cage. The remainder of the internal aspect of the cage is covered with mulch and potted plants to emulate a typical yard. An air conditioning system comprised of two external ACs that feed cooled, moistened air into the cage units. The air is released from the central ceiling beam from a long cloth tube that disperses the airflow and also prevents mosquitoes from escaping the cage via the AC system. Sensors located inside and outside the cage monitor ambient temperature and relative humidity, with AC controlled to match ambient conditions. Data loggers set in the cages and outside found a <2°C temperature difference. Additional security features include air curtains over exit doors, sticky traps to monitor for escaping mosquitoes between layers of the mesh, a lockable vestibule leading from the connecting corridor to the cage and from inside to outside of the insectary, and screened (0.25 mm mesh) drains within the insectary and the cage. A set of standard operating procedures (SOP) has been developed to ensure that security is maintained and for enhanced surveillance for escaping mosquitoes on the JCU campus where the MRF is located. A cohort of male and female Aedes aegypti mosquitoes were released in the cage and sampled every 3–4 days to determine daily survival within the cage; log linear regression from BG-sentinel trapping collections produced an estimated daily survival of 0.93 and 0.78 for females and males, respectively.

Conclusions/Significance

The MRF SFS allows us to test novel control strategies within a secure, contained environment. The air-conditioning system maintains conditions within the MRF cages comparable to outside ambient conditions. This cage provides a realistic transitional platform between the laboratory and the field in which to test novel control measures on quarantine level insects.     

Male size and mating success in Drosophila melanogaster and D. pseudoobscura under field conditions

Wild mating male D. melanogaster and D. pseudoobscura were larger than randomly sampled males. That this was due to size and not a confounding effect of age was confirmed by release experiments with D. melanogaster of standard age; larger males were at a mating advantage with virgin and inseminated females. In both species larger males delivered more courtship and in D. pseudoobscura they won more aggressive encounters. These results on Drosophila in the field confirm and extend those previously obtained in the laboratory.     

The impact of insecticide‐treated cloth targets on the survival of Stegomyia polynesiensis (= Aedes polynesiensis) under laboratory and semi‐field conditions in French Polynesia

The impact of deltamethrin‐impregnated cloth targets on Stegomyia polynesiensis (= Aedes polynesiensis) (Marks) (Diptera: Culicidae) was assessed under laboratory and semi‐field settings in French Polynesia. Stegomyia polynesiensis females were released into small laboratory cages and large field cages containing either a deltamethrin‐treated or an untreated navy blue cloth, and mosquito knock‐down and mortality were assessed. The 24‐h mortality rate in mosquitoes exposed to the insecticide‐treated target in small cages was 98.0%. These mosquitoes also demonstrated significantly higher levels of knock‐down than those exposed to the untreated target. Mortality in field cages was assessed at 24 and 48 h. The 24‐h mortality rate in mosquitoes exposed to the control target was 31.2%, whereas that in those exposed to the deltamethrin‐treated target was 54.3%. The 48‐h mortality rate was also elevated in mosquitoes exposed to the deltamethrin‐treated target, but this result did not differ significantly from that observed in mosquitoes exposed to the control target. The significant suppression of female S. polynesiensis by deltamethrin‐treated resting targets in this study indicates that these targets could play a role in the control of an important disease vector in the South Pacific region.     

Genistein preincubation of Bradyrhizobium japonicum cells improves strain competitiveness under greenhouse,but not field conditions

In the soybean-B. japonicum symbiosis, genistein has been identified as one of the major compounds in soybean seed and root exudates responsible for inducing expression of the B. japonicum nodYABC operon. In this study, we have tested the possibility that genistein treatment prior to inoculation can increase the competitiveness of the treated B. japonicum strain under both greenhouse and field conditions. Two mutants of the two B. japonicum strains each with a different antibiotic resistant marker were selected. They were tested with one or the other treated with genistein. The results showed genistein treated mutants had higher levels of nodule occupancy than the untreated mutant or parent strain under greenhouse conditions. Mutants from 532C had higher nodule occupancies than mutants from USDA110, especially at 15 °C. In the more complex field environment, genistein treated mutants formed fewer nodules than the untreated mutants. The contradictory results of strain competitiveness for greenhouse and field experiments are discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

Landing response of Aedes (Stegomyia) polynesiensis mosquitoes to coloured targets

Aedes polynesiensis Marks (Diptera: Culicidae) is the primary vector of lymphatic filariasis (LF) in the island countries and territories of the South Pacific. In the development of a novel control tool, the response of Ae. polynesiensis to six different colours (three solid fabrics, two patterned fabrics and a plastic tarp) was measured using a digital photographic system. Adult mosquitoes were placed into an environmental chamber and allowed to choose between a white target and one of six experimental targets. Mosquito landing frequency and landing duration were calculated. Adult female Ae. polynesiensis preferred all of the experimental targets to the white control target. Mosquito landing frequency was highest for the solid targets (black, navy blue and red) followed in turn by the two colour pattern targets and the polyethylene target. Mosquito landing duration was greater for experimental targets when compared with white control targets. Mosquito landing frequencies did not change over time during the course of the assay. The response of male Ae. polynesiensis was also measured when exposed to a 100% cotton black target. Male mosquitoes preferred the black target to the white control target, although at levels lower than that observed in female mosquitoes. The results suggest that future investigations evaluating the visual responses of Ae. polynesiensis mosquitoes are warranted, with a special emphasis on semi‐field and field‐based experiments.     

Aedes polynesiensis in the Society Islands: environmental correlates of isoenzyme differentiation

Abstract Isoenzyme genetic differentiation of Aedes polynesiensis mosquitoes in Raiatea island, French Polynesia, was evaluated by two models of population structure based on seven gene-enzyme systems: Ak, Est, Got, Gpi, Hk, Mdh and Pgm. The ecological model tested whether genetic differentiation is congruent with habitat differences. The isolation model evaluated whether genetic differentiation is proportional to geographical distribution. The ecological model found no significant differentiation between populations of Ae.polynesiensis from beach and forest ecotopes, whereas the isolation model was consistent with the data. However, incipient speciation is opposed by the source-sink system of population dynamics in such small neighbouring islands, where Ae.polynensiensis extinction is readily followed by reinvasion causing considerable gene flow between island populations.     

The presence of knockdown resistance mutations reduces male mating competitiveness in the major arbovirus vector,Aedes aegypti

BackgroundThe development of insecticide resistance in mosquitoes can have pleiotropic effects on key behaviours such as mating competition and host-location. Documenting these effects is crucial for understanding the dynamics and costs of insecticide resistance and may give researchers an evidence base for promoting vector control programs that aim to restore or conserve insecticide susceptibility.Methods and findingsWe evaluated changes in behaviour in a backcrossed strain of Aedes aegypti, homozygous for two knockdown resistance (kdr) mutations (V1016G and S989P) isolated in an otherwise fully susceptible genetic background. We compared biting activity, host location behaviours, wing beat frequency (WBF) and mating competition between the backcrossed strain, and the fully susceptible and resistant parental strains from which it was derived. The presence of the homozygous kdr mutations did not have significant effects on blood avidity, the time to locate a host, or WBF in females. There was, however, a significant reduction in mean WBF in males and a significant reduction in estimated male mating success (17.3%), associated with the isolated kdr genotype.ConclusionsOur results demonstrate a cost of insecticide resistance associated with an isolated kdr genotype and manifest as a reduction in male mating success. While there was no recorded difference in WBF between the females of our strains, the significant reduction in male WBF recorded in our backcrossed strain might contribute to mate-recognition and mating disruption. These consequences of resistance evolution, especially when combined with other pleiotropic fitness costs that have been previously described, may encourage reversion to susceptibility in the absence of insecticide selection pressures. This offers justification for the implementation of insecticide resistance management strategies based on the rotation or alternation of different insecticide classes in space and time.     

Productivity of natural and artificial containers for Aedes polynesiensis and Aedes aegypti in four American Samoan villages

Six mosquito species were identified in a survey of containers associated with 347 households in four villages in American Samoa. Aedes polynesiensis Marks (Diptera: Culicidae) and Aedes aegypti (L) were the most abundant species, representing 57% and 29% of the mosquitoes identified. Culex quinquefasciatus (Say), Culex annulirostris (Skuse), Aedes oceanicus (Belkin) and Toxorhynchites amboinensis (Doleschall) were also found. Aedes aegypti and Ae. polynesiensis showed distinct differences in their use of containers, preferring large and small containers, respectively. By contrast with previous studies, Ae. polynesiensis utilized domestic and natural containers with equal frequency, whereas Ae. aegypti continued to be found predominantly in domestic containers. Only 15% of containers holding immature mosquitoes included pupae and fewer than 10 Aedes spp. pupae were found in most containers with pupae. An estimated 2289 Ae. polynesiensis and 1640 Ae. aegypti pupae were found in 2258 containers. The presence of both species in the same container did not affect the mean density of either species for larvae or pupae. Glass jars, leaf axils, tree holes and seashells produced few Aedes spp. pupae in any of the study villages. Overall, 75% of Ae. polynesiensis pupae were found in buckets, ice-cream containers and tyres, with <7% being produced in natural containers, whereas 82% of Ae. aegypti pupae were found in 44-gallon (US) drums ( approximately 166L), buckets and tyres. Source reduction efforts targeting these container types may yield significant reductions in both Ae. polynesiensis and Ae. aegypti populations in American Samoa.     

Survival of colour-polymorphic Eristalis arbustorum hoverflies in semi-field conditions

1. The theory on adaptive phenotypic plasticity assumes different fitness optima in different environments. To demonstrate these fitness differences in the field or under laboratory conditions can be difficult.
2. The size of the coloured patches on the abdomen of Eristalis arbustorum varies with rearing temperature. At low temperatures small patches develop and at higher temperatures larger patches develop. Consequently, the colour patch size varies seasonally.
3. To study the relative fitness of two extreme phenotypes in abdominal coloration in the summer, a total of 591 laboratory-reared flies were released in six large population cages. As a measure of fitness, survival was estimated.
4. In all replicate cages the survival of the pale phenotypes was higher than that of the dark phenotypes, indicating a higher fitness for the pale flies under summer conditions.
5. The fat content of flies reared under identical conditions to those released in the cages was measured. Pale flies had ≈ 0·7 mg more fat than dark flies (± 25% extra fat). However, it is argued that this difference in fat reserves could not have been the only cause of the observed difference in survival.
6. The results are discussed in relation to mimicry and thermoregulation.     

Female-biased dispersal under conditions of low male mating competition in a polygynous mammal

Sex-biased dispersal is a common phenomenon in birds and mammals. Competition for mates has been argued to be an important selective pressure favouring dispersal. Sexual differences in the level of intrasexual competition may produce asymmetries in the costs-benefits balance of dispersal and philopatry for males and females, which may favour male-biased dispersal in polygynous species such as most mammals. This being the case, condition-dependent dispersal predicts that male-bias should decrease if mating competition relaxes. We test this expectation for red deer, where male-biased dispersal is the norm. In southwestern Spain, red deer populations located in nonfenced hunting estates presented altered structures with sex ratio strongly biased to females and high proportion of young males. As a consequence, mate competition in these populations was lower than in other, most typical red deer populations. We found that, under such conditions of altered population structure, dispersal was female-biased rather than male-biased. Additionally, mate competition positively related to male dispersal but negatively to female dispersal. Other factors such as resource competition, age of individuals and sex ratio were not related to male or female dispersal. Males may not disperse if intrasexual competition is low and then females may disperse as a response to male philopatry. We propose hypotheses related to female mate choice to explain female dispersal under male philopatry. The shift of the sex-biased dispersal pattern along the gradient of mate competition highlights its condition-dependence as well as the interaction between male and female dispersal in the evolution of sex-biased dispersal.     

Performance of transgenic spring wheat plants and effects on non-target organisms under glasshouse and semi-field conditions

Abstract:  A convertible glasshouse was established to study annual transgenic plants under near-field environmental conditions while simultaneously ensuring a high level of biological containment. This system can provide a useful step in the assessment of transgenic plants prior to open-field experiments. Two transgenic wheat lines (cv. Bobwhite) were investigated and compared with their corresponding non-transformed wildtypes with respect to plant performance, expression of the transgenic trait and interactions with antagonists. The first line expressed snowdrop lectin [ Galanthus nivalis agglutinin (GNA)] for enhanced resistance to aphids, and the second one overexpressed the endogenous Lr10 gene to enhance resistance to leaf rust. Interestingly, 1000-kernel weight of Lr10 -transgenic plants was significantly reduced, indicating that the overexpression of the Lr10 gene caused a significant fitness cost for the plant. GNA-transgenic plants expressed the lectin at levels too low to affect the target aphids. A detached leaf bioassay with Lr10 -transgenic plants revealed an increased resistance to leaf rust. No differences in the performance of aphids or cereal leaf beetles on transgenic and non-transformed plants were recorded in the convertible glasshouse and in complementary glasshouse studies. Similarly, infection levels with powdery mildew did not differ between transgenic and non-transformed plants but Bobwhite plants were significantly more infected when compared with conventional Swiss spring wheat cultivars. Overall, the assessment revealed that for the plants investigated here, their genetic background had a stronger impact on the performance of a plant and its interactions with insect herbivores and pathogens than the expression of the transgene.     

Male mating performance and cytoplasmic incompatibility in a wPip Wolbachia trans‐infected line of Aedes albopictus (Stegomyia albopicta)

Wolbachia pipientis Hertig (Rickettsiales: Rickettsiaceae) is a maternally inherited endosymbiont of a large number of insects and other arthropods that induces various effects on host reproductive biology. Among these, cytoplasmic incompatibility (CI) is a form of sterility induced in eggs produced by mating between infected males and females uninfected or infected by an incompatible Wolbachia strain. This phenomenon has been proposed as a potential way to produce functionally sterile males to be used in genetic control programmes. In this paper, we report on experiments carried out to evaluate the mating performances of males of an Aedes albopictus (Stegomyia albopicta) (Diptera: Culicidae) line (ARwP), harbouring a new Wolbachia infection [the wPip strain from Culex pipiens Linnaeus (Diptera: Culicidae)], in comparison with naturally infected males (SR line). ARwP males did not differ from SR males with regard to insemination capacity. Mating competitiveness did not differ significantly between lines in either laboratory or greenhouse conditions. Moreover, crosses with SR females were characterized by a 100% CI regardless of ARwP male age. All of these findings suggest that ARwP males may represent a very efficient tool for control programmes against Ae. albopictus based on the release of functionally sterile males.     

Male mating strategies and the mating system of great-tailed grackles

Great-tailed grackles (Quiscalus mexicanus) are sexually dimorphic,dichromatic, colonially nesting blackbirds. In this study, males pursued three basic types of conditional mating strategies,each of which employed a different set of mating tactics. Territorialmales defended one or more trees in which several females nested.They achieved reproductive success by siring the offspringof their social mates and through extrapair fertilization.Resident males lived in the colony but did not defend territoriesor have social mates. Transient males passed through the colony, staying no more than a few days, and probably visited more thanone colony. Residents appeared to queue for access to territories,but transients did not. Residents and transients gained allpaternity through extrapair fertilizations and provided noparental care. Territorial males sired the majority of offspring,but residents and transients also sired small numbers of nestlings. Territorial males were larger and had longer tails than nonterritorialmales. The number of social mates was related to body size,and males that sired nestlings were heavier and had longertails than males with no genetic reproductive success. Malesthat gained paternity through extrapair fertilization wereheavier and had longer tails than males that did not. The matingsystem of great-tailed grackles can best be categorized as "non-faithful-female frank polygyny."     

Acoustic orientation of male Aedes diantaeus during mating

The reaction of males of Aedes diantaeus to different auditory stimuli was studied in natural environment during swarming and under laboratory conditions. The males were attracted by 250-420 Hz that corresponds to the main frequencies of flight tones of sympatric females and the range of sensitivity of male Johnston's organs. Behavioural data show that in natural environment swarming males are attracted by flying conspecific and heterospecific females rather than by dead or immobilized females. Contact with heterospecific females always ended in parting of mosquitoes while contact with cospecific females resulted in pairing. Thus, swarming males of Aedes diantaeus localize females by their flight tones but recognize the females of their own species in contact.     

The relative importance and distribution of Aedes polynesiensis and Ae.aegypti larval habitats in Samoa

Abstract. In preparation for a Filariasis Control programme in Samoa, during 1978 monthly larval surveys of the vector mosquito Aedes polynesiensis were carried out in four study villages in the main island of Upolu. A more extensive survey of larval habitat distribution was then made in twenty-two villages of Upolu and eighteen of Savai'i island, to determine the importance of habitat types according to their abundance, volume of water and whether their productivity was permanent or seasonal. Ae.aegypti larval densities and habitat distribution were also monitored and the occurrence of predatory Toxorhynchites amboinensis larvae in northern Upolu was recorded from forty-one collections. Aedes Breteau and container indices fluctuated with the pattern of rainfall in two coastal villages and an inland bush village, but not in a coconut plantation community. The five main Aedes larval habitat types encountered were: 200 litre water-storage drums, discarded tins and bottles, coconut shells, automobile tyres and treeholes. Aedes immatures occurred perennially in drums and tree holes, but breeding discontinued in tins, bottles and coconut shells during the driest month of July. For Ae.polynesiensis in Upolu the Breteau and container indices of 104.5 ± SD 80.9 and 35.3 ± 12.4 respectively were significantly higher than those in Savai'i: 33.1 ±25.0 and 24.3 ±20.0 respectively. Likewise for Ae.aegypti the Breteau and container indices of 50.8 ± 32.5 and 23.9 ± 15.6, respectively, were also significantly higher than those in Savai'i: 12.7 ± 17.1 and 9.4 ±13.2 respectively. Habitat types greater or lesser importance were determined by plotting the percentage of each type of container utilized for Aedes breeding against the percentage of each type amongst all larva-positive containers. Ae.polynesiensis preferred tree-holes but not water-storage drums. Ae.aegypti preferred drums and tyres; mixed populations of larvae of both species were commonest in these two types of habitat. Ae.polynesiensis occurred in every village. Ae.aegypti was encountered in all twenty-two villages surveyed in Upolu and nine of eighteen villages in Savai'i. Total larval surveys revealed that drums and tree-holes contained the highest numbers of Aedes larvae. The study provided criteria for planning a control programme.     

Colonization of a hybrid strain to restore male Anastrepha ludens (Diptera: Tephritidae) mating competitiveness for sterile insect technique programs

To restore male mating competitiveness of Mexican fruit flies, Anastrepha ludens (Loew) (Diptera: Tephritidae), reared for sterile insect releases by the Mexican Fruit fly Eradication Campaign, two strain replacement techniques were evaluated. Field cage male competitiveness tests revealed that laboratory males of the Metapa strain mated 3 times less often with wild females than field-collected wild males. A strain developed from the cross of wild males and laboratory females (hybrid strain) was similar to a strain developed from the cross of laboratory males and females (laboratory strain) in that its females produced similar amounts of eggs and the eggs displayed similar levels of hatch and egg-to-pupa transformation in artificial diet. By contrast, a strain developed from the cross of wild males and females (wild strain), forced into artificial rearing, experienced a series of bottlenecks involving reduced egg laying and extremely poor development in diet. The male F1 progeny of the hybrid strain and field-collected wild males outcompeted Fl laboratory males in field cage tests for matings with field-collected wild females. In conclusion, we found that strains developed from the cross of wild males and laboratory females allowed us to restore male mating competitiveness of F1 Mexican fruit flies without compromising mass-rearing production.