A new sex-linked mutant short wing in Aedes aegypti.

A spontaneous recessive sex-linked mutant short wing has been discovered in the mosquito Aedes aegypti. It is situated less than one cross-over unit from the sex determining locus. In homozygous females, flight is impaired and the survival and fecundity is markedly subnormal. Two possible uses of this gene for genetic control operations are envisaged: (a) to provide automatic sexing of males for release and (b) enhancement of the population control potential of other available genetics systems.     

Trisomy in Aedes aegypti.

A trisomic (2n=6+1) pupa of the yellow fever mosquito Aedes aegypti has been found. The trisomy involved chromosome 3 which is intermediate in size between 1 and 2. The extra chromosome formed a univalent or a trivalent during meiosis.     

Dominant-and-recessive epistasis in a homeotic mosquito mutant.

Following selection for 15 generations a pure strain of a homeotic mutant spur was isolated from a Brazilian population of the mosquito Culex pipiens fatigans. Monohybrid crosses showed a 13:3 segregation indicating dominant-and-recessive epistasis for wild-type vs. spur. This implies that a dominant allele at one locus and a recessive at the other interact to produce the mutant phenotype. Dihybrid crosses with linkage group II markers yellow and ruby gave 39:13:9:3 ratios indicating independent segregation. However, the dihybrid cross with linkage group I marker maroon showed a highly significant departure from 39:13:9:3 ratio. Data available indicate that the phenotype spur is controlled by a dominant epistat in linkage group III and a recessive epistat (approximately 31.9 crossover units from maroon) in linkage group I.     

Aedes aegypti ferritin.

Diseases transmitted by hematophagous (blood-feeding) insects are responsible for millions of human deaths worldwide. In hematophagous insects, the blood meal is important for regulating egg maturation. Although a high concentration of iron is toxic for most organisms, hematophagous insects seem unaffected by the iron load in a blood meal. One means by which hematophagous insects handle this iron load is, perhaps, by the expression of iron-binding proteins, specifically the iron storage protein ferritin. In vertebrates, ferritin is an oligomer composed of two types of subunits called heavy and light chains, and is part of the constitutive antioxidant response. Previously, we found that the insect midgut, a main site of iron load, is also a primary site of ferritin expression and that, in the yellow fever mosquito, Aedes aegypti, the expression of the ferritin heavy-chain homologue (HCH) is induced following blood feeding. We now show that the expression of the Aedes ferritin light-chain homologue (LCH) is also induced with blood-feeding, and that the genes of the LCH and HCH are tightly clustered. mRNA levels for both LCH- and HCH-genes increase with iron, H2O2 and hemin treatment, and the temporal expression of the genes is very similar. These results confirm that ferritin could serve as the cytotoxic protector in mosquitoes against the oxidative challenge of the bloodmeal. Finally, although the Aedes LCH has no iron responsive element (IRE) at its 5'-untranslated region (UTR), the 5'-UTR contains several introns that are alternatively spliced, and this alternative splicing event is different from any ferritin message seen to date.     

Highly-substrate active isoenzyme acetylcholinesterase-II,in rosy eye mutant of Aedes aegypti mosquito

Insecticide bioassays were carried out on larvae and adults of rosy eye mutant and wildtype strains of A. aegypti. Both the strains were equally susceptible to DDT, malathion and deltamethrin. Biochemical assays showed an increase in acetylcholinesterase enzyme (AChE) activity in all the stages of mutant strain with both the substrates i.e. acetylthiocholine iodide and S-butyrylthiocholine iodide. However, there was no difference in the percent inhibition of enzyme activity with propoxur in these two strains. Polyacrylamide gel electrophoresis performed in native conditions on the homogenates of adults of rosy eye mosquitoes showed that AChE-II allele was highly active with the substrate acetylthiocholine iodide as compared to wildtype strain. Frequency of the highly active AChE-II allele in the mutant strain was about 68%, whereas it was about 5% in the wildtype strain.     

FLP-mediated recombination in the vector mosquito, Aedes aegypti.

The activity of a yeast recombinase, FLP, on specific target DNA sequences, FRT, has been demonstrated in embryos of the vector mosquito, Aedes aegypti. In a series of experiments, plasmids containing the FLP recombinase under control of a heterologous heat-shock gene promoter were co-injected with target plasmids containing FRT sites into preblastoderm stage mosquito embryos. FLP-mediated recombination was detected between (i) tandem repeats of FRT sites leading to the excision of specific DNA sequences and (ii) FRT sites located on separate plasmids resulting in the formation of heterodimeric or higher order multimeric plasmids. In addition to FRT sites originally isolated from the yeast 2 microns plasmid, a number of synthetic FRT sites were also used. The synthetic sites were fully functional as target sites for recombination and gave results similar to those derived from the yeast 2 microns plasmid. This successful demonstration of yeast FLP recombinase activity in the mosquito embryo suggests a possible future application of this system in establishing transformed lines of mosquitoes for use in vector control strategies and basic studies.     

Aedes aegypti genomics

The mosquito, Aedes aegypti, is the primary, worldwide arthropod vector for the yellow fever and dengue viruses. As it is also one of the most tractable mosquito species for laboratory studies, it has been and remains one of the most intensively studied arthropod species. This has resulted in the development of detailed genetic and physical maps for Ae. aegypti and considerable insight into its genome organization. The research community is well-advanced in developing important molecular tools that will facilitate a whole genome sequencing effort. This includes generation of BAC clone end sequences, physical mapping of selected BAC clones and generation of EST sequences. Whole genome sequence information for Ae. aegypti will provide important insight into mosquito chromosome evolution and allow for the identification of genes and gene function. These functions may be common to all mosquitoes or perhaps unique to individual species, possibly specific to host-seeking and blood-feeding behaviors, as well as the innate immune response to pathogens encountered during blood-feeding. This information will be invaluable to the global effort to develop novel strategies for preventing arthropod-borne disease transmission.     

Ammonia metabolism in Aedes aegypti

We investigated the mechanisms by which Aedes aegypti mosquitoes are able to metabolize ammonia. When females were given access to solutions containing NH(4)Cl or to a blood meal, hemolymph glutamine and proline concentrations increased markedly, indicating that ammonium/ammonia can be removed from the body through the synthesis of these two amino acids. The importance of glutamine synthetase was shown when an inhibitor of the enzyme was added to the meal causing the glutamine concentration in hemolymph to decrease significantly, while the proline concentration increased dramatically. Unexpectedly, we found an important role for glutamate synthase. When mosquitoes were fed azaserine, an inhibitor of glutamate synthase, the glutamine concentration increased and the proline concentration decreased significantly. This confirms the presence of glutamate synthase in mosquitoes and suggests that this enzyme contributes to the production of glutamate for proline synthesis. Several key enzymes related to ammonium/ammonia metabolism showed activity in homogenates of mosquito fat body and midgut. The mosquito genes encoding glutamate dehydrogenase, glutamine synthetase, glutamate synthase, pyrroline-5-carboxylate synthase were cloned and sequenced. The mRNA expression patterns of these genes were examined by a real-time RT-PCR in fat body and midgut. The results show that female mosquitoes have evolved efficient mechanisms to detoxify large loads of ammonium/ammonia.     

Gustatory stimulation of a homeotic mutant appendage,Antennapedia, inDrosophila melanogaster

Summary The proboscis extension response was used to prove the leg identity of chemosensory neurons in the homeotic appendage of theDrosophila mutantAntennapedia (Antp ^73b). The data suggest that the homeotic appendage, which is morphologically characterized as a mesothoracic leg, corresponds to a mesothoracic leg as well when considering its gustatory responsiveness (Figs. 1A, B; 3A, B). The neuronal pathway which might mediate the reflex between homeotic chemoreceptors and motor neurons responsible for the proboscis extension is discussed.     

Neutral lipids of Aedes aegypti and Aedes albopictus cells cultured in vitro.

Aedes aegypti feeding on chickens infected with Plasmodium gallinaceum take less blood and lay fewer eggs than those feeding on uninfected hosts. Both activities show an inverse correlation with the degree of parasitemia. Mosquitoes feeding on infected chickens ingest blood in amounts directly proportional to the length of time spent on the hosts, whereas there is no relationship between host contact and blood meal size for mosquitoes feeding on uninfected hosts. Feeding and probing choice experiments demonstrate that infected chickens are less attractive to Aedes aegypti than uninfected chickens.     

Stamenless, a tomato mutant with homeotic conversions in petals and stamens

A tomato (Lycopersicon esculentum Mill.) monogenic semidominant mutation, stamenless (sl), which results in homeotic conversions in two adjacent floral whorls, was studied. When grown at standard temperature, flowers of sl/sl plants showed sepaloid petals in the second whorl and strong transformation of stamens to carpels in whorl three. These transformed carpels were fused with each other and with the genuine carpels in the fourth whorl to form a unique gynoecium. The mutation is semidominant since heterozygous plants showed a phenotype intermediate between that of the wild type (WT) and that of homozygous mutant plants, with nearly WT petals but with feminized stamens bearing naked ovules on the base of their adaxial face. The initiation and position of organ primordia in sl/sl flowers were not altered when compared with WT primordia although development of organ primordia in the second and third whorls deviated from WT at an early stage as observed by scanning electron microscopy. The mutant phenotype is temperature sensitive and when sl/sl plants were cultured at low temperature, the morphology of some flowers resembled that of the WT. This reversion of the mutant phenotype is also induced by treatment of young sl/sl plants with gibberellic acid, providing evidence that gibberellin synthesis or sensitivity could mediate the effect of low temperature on the mutant phenotype. Southern blot analyses using a Deficiens-homologous gene from Solanum tuberosum as a probe showed a restriction-fragment-length polymorphism (RFLP) linked to the sl mutation. This result indicates that the mutation affects a Deficiens-like gene that controls the identity of petals and stamens. Received: 10 December 1998 / Accepted: 29 March 1999     

Herpetomonas,with Bacterium-like Inclusions,in Malaysian Aedes aegypti and Aedes albopictus*

SYNOPSIS Herpetomonas sp. was found repeatedly in the Malpighian tubules of laboratory-reared male and female Aedes aegypti and Aedes albopictus in Malaysia. The flagellates occurred irregularly, in batches, and were absent during long periods. The data suggest an exogenous source of infection for the mosquitoes, presumably from another insect, probably of another genus. Thirty to 40% of the flagellates of Aedes contained intracytoplasmic rod-shaped structures strongly resembling bacteria. These were found often in groups suggesting intracellular multiplication. They were passed to the Herpetomonas daughter cells during division. Parasitism of Aedes by Herpetomonas is extremely unusual, only one previous record, an inconclusive one, having been found. Parasitism by Herpetomonas containing bacterium-like rods has apparently never been reported.     

Food as a limiting factor for Aedes aegypti in water-storage containers.

An understanding of the ecological factors that regulate natural populations of Aedes aegypti mosquitoes can improve control and reduce the incidence of dengue (DF) and dengue hemorrhagic fever (DHF) in tropical areas. We investigated whether immature Ae. aegypti in water-storage containers from an urban area were under food limitation. We used starvation resistance (number of days alive without food) as an indicator of the feeding history in third-instar Ae. aegypti larvae. Resistance to starvation and other measures of immature success, such as development time, survival, and adult mass, were investigated across a wide range of feeding conditions in the laboratory. Resistance to starvation of third-instar larvae and body mass of adults emerging from pupae collected in water-storage containers in an urban area were compared with the laboratory results. If resistance to starvation and adult mass of field-collected Ae. aegypti corresponded with the lower levels of feeding in the laboratory, then food limitation could be inferred in field-collected larvae. Results showed that resistance to starvation was well correlated with previous feeding levels and with the other measures of immature success. Both resistance to starvation and adult body mass of field-collected specimens corresponded with the lower levels of feeding in the laboratory. Therefore, it was concluded that food limitation or competition is likely to be a regulatory factor in water-storage containers in the urban area. It is recommended that any control measure applied to immature Ae. aegypti in water-storage containers should eliminate all or most of the individuals, otherwise unintended, undesirable results might occur, such as the production of more and larger adults.     

Midgut bacterial dynamics in Aedes aegypti

In vector mosquitoes, the presence of midgut bacteria may affect the ability to transmit pathogens. We have used a laboratory colony of Aedes aegypti as a model for bacterial interspecies competition and show that after a blood meal, the number of species (culturable on Luria-Bertani agar) that coexist in the midgut is low and that about 40% of the females do not harbor any cultivable bacteria. We isolated species belonging to the genera Bacillus, Elizabethkingia, Enterococcus, Klebsiella, Pantoea, Serratia, and Sphingomonas, and we also determined their growth rates, antibiotic resistance, and ex vivo inhibition of each other. To investigate the possible existence of coadaptation between midgut bacteria and their host, we fed Ae.?aegypti cohorts with gut bacteria from human, a frog, and two mosquito species and followed the bacterial population growth over time. The dynamics of the different species suggests coadaptation between host and bacteria, and interestingly, we found that Pantoea stewartii isolated from Ae.?aegypti survive better in Ae.?aegypti as compared to P.?stewartii isolated from the malaria mosquito Anopheles gambiae.     

Distention-mediated egg maturation in the mosquito,Aedes aegypti

Abdominal distention accelerates the release of a factor from the head of blood-fed Aedes aegypti mosquitoes. The critical period during which the head is required for oögenesis following blood ingestion is approx 6 h with a 5 μl meal, but small blood meals of 1 μl require the head to be present for significantly longer. Increasing the abdominal distention by supplementing the 1 μl meal with saline results in a critical period similar to that with 5 μl of blood. The information from the distended abdomen appears to travel via the ventral nerve cord. Transection of the ventral nerve cord prevents oögenesis from occurring after small blood meals, but not with larger blood volumes. Topical application of 100 pg of juvenile hormone III can substitute for the distention message.