A behavioural and electrophysiological study of ovi position cues for Culex quinquefasciatus

Abstract. Both the synthetic oviposition pheromone, erythro-6-acetoxy-5-hexa-decanolide (0.01–80 μg) and a polluted water sample (0.01–80%) significantly increased oviposition by gravid female Culex quinquefasciatus Say (Diptera: Culicidae) in a laboratory assay against clean water. An additive effect resulted when 0.05 μg oviposition pheromone was combined with the polluted water dilution series. Undiluted, however, both the oviposition pheromone and the polluted water sample, alone and in combination, reduced the oviposition response to control levels. A previously reported oviposition site odour component, 3-methylindole, also significantly increased oviposition at 1 × 10^_8 to 1 × 10^-7g but at 1 × 10^_7g significantly less oviposition occurred than in control treatments. The biological activities of the compounds were additionally evaluated electrophysiologically by an EAG assay. Female mosquitoes were electrophysiologically sensitive to the oviposition cues, both pheromone and habitat-related; EAG responses of males were considerably lower than of females with 3-methylindole.     

Silent,generic and plant kairomone sensitive odorant receptors from the Southern house mosquito

The Southern house mosquito Culex quinquefasciatus has the largest repertoire of odorant receptors (ORs) of all mosquitoes and dipteran species whose genomes have been sequenced to date. Previously, we have identified and de-orphanized two ORs expressed in female antennae, CquiOR2 and CquiOR10, which are sensitive to oviposition attractants. In view of a new nomenclature for the Culex genome (VectorBase) we renamed these ORs as CquiOR21 (formerly CquiOR10) and CquiOR121 (CquiOR2). In addition, we selected ORs from six different phylogenetic groups for deorphanization. We cloned four of them by using cDNA from female antennae as a template. Attempts to clone CquiOR87 and CquiOR110 were unsuccessful either because they are pseudogenes or are not expressed in adult female antennae, the main olfactory tissue. By contrast, CquiOR1, CquiOR44, CquiOR73, and CquiOR161 were highly expressed in female antennae. To de-orphanize these ORs, we employed the Xenopus oocyte recording system. CquiORx–CquiOrco-expressed oocytes were challenged with a panel of 90 compounds, including known oviposition attractants, human and vertebrate host odorants, plant kairomones, and naturally occurring repellents. While CquiOR161 did not respond to any test compound in two different laboratories, CquiOR1 showed the features of a generic OR, with strong responses to 1-octen-3-ol and other ligands. CquiOR44 and CquiOR73 showed preference to plant-derived terpenoids and phenolic compounds, respectively. While fenchone was the best ligand for the former, 3,5-dimethylphenol elicited the strongest responses in the latter. The newly de-orphanized ORs may be involved in reception of plant kairomones and/or natural repellents.     

Development and laboratory evaluation of chemically‐based baited ovitrap for the monitoring of Aedes aegypti

Aedes (Stegomyia) aegypti is considered to be the most important dengue vector worldwide. Studies were conducted to design and evaluate a chemically‐based baited ovitrap for monitoring Ae. aegypti under laboratory conditions. Several known chemical attractants and three types of ovitraps (ovitraps A, B, and C) were evaluated throughout the oviposition bioassays. Oviposition responses of gravid female Ae. aegypti were evaluated to n‐heneicosane, 3‐methylindole (skatole), 4‐methylphenol (p‐cresol), and phenol. Female Ae. aegypti were attracted to all the evaluated compounds. Among them, n‐heneicosane at a concentration of 10 ppm (mg/l), skatole from 50 to 1000 ppm, p‐cresol at 100 ppm, and phenol at 50 ppm showed a significant positive oviposition response. A blend of the four chemical attractants increased the oviposition response; 67% of the eggs were deposited in the treatment compared to the control. Female Ae. aegypti were signi?cantly more attracted to ovitrap A loaded with the four‐component synthetic blend compared to the standard ovitrap in the oviposition bioassays. The compound used in ovitrap A retained its attractant property for up to three days. The chemically‐based baited ovitrap may be considered as an option to be integrated during the monitoring of dengue virus vectors in México.     

Cardiac glycosides and oviposition by Danaus plexippus on Asclepias fruticosa in south-east Queensland (Australia), with notes on the effect of plant nitrogen content

Abstract. 1. Previous studies which attempt to relate oviposition by female monarchs, Danaus plexippus (L.), to the cardiac glycosides of their milkweed host plants are reviewed and found to be defective.
2. Here we followed ovipositing females in the field and recorded their oviposition behaviour on individual Asclepias fruticosa (L.) plants. The cardenolide glycoside and per cent nitrogen of each of these plants and also of a control set of plants was recorded.
3. Milkweeds accepted by females, on which egg laying took place, had significantly lower mean cardenolide levels compared with rejected plants and a control sets of plants.
4. Most oviposition ( c. 70%) took place on plants with intermediate cardenolide (CG) levels (=200–300 μg/0.1 g DW).
5. The mean CG level in A.fruticosa was 345 μg/0.1 g DW (SD = 138, n =122, range 119–719).
6. There was no relationship between monarch oviposition behaviour and plant per cent nitrogen.     

Laboratory tests of the effects of p-cresol and 4-methylcyclohexanol on oviposition by three species of Toxorhynchites mosquitoes

Laboratory experiments tested the effects of p-cresol or 4-methylcyclohexanol at concentrations of 1, 10 and 50 ppm, on oviposition by the mosquitoes Toxorhynchites brevipalpis Theobald, Tx. amboinensis (Doleschall) and Tx. splendens (Wiedemann). A 5 + 5 ppm mixture of the two chemicals was also tested. All three species laid significantly more eggs in cups containing p-cresol, whereas only Tx. brevipalpis and Tx. amboinensis responded similarly to 4-methylcycohexonol and to the mixture of both chemicals. Tx. brevipalpis was, to a relatively limited degree, the most responsive of the three species. Ancillary experiments indicated that the chemicals were acting as attractants, causing more females to fly to treated cups. No stimulant effects were detected either in terms of the proportion of females that initiated oviposition flight (after flying to the cups) or in terms of the number of looping flights executed prior to ejection of an egg.     

Electrophysiological and behavioural responses of Scyphophorus acupunctatus (Col., Curculionidae) to Agave tequilana volatiles

Abstract:  The weevil Scyphophorus acupunctatus Gyllenhal (Col., Curculionidae) is the most important insect pest of cultivated agaves in Mexico. The objective of this study was to search for potential attractants for this weevil species from the freshly cut fleshy leaves of healthy Agave tequilana Weber var. Blue plants. Combined gas chromatography-electroantennography (GC-EAD) analysis of A. tequilana volatile extracts showed that seven peaks elicited antennal responses from males and females. Five of these peaks were identified by gas chromatography-mass spectrometry (GC-MS) as o -xylene, α -pinene, 3-carene, γ -terpinene and linalool. Electroantennogram (EAG) records of both sexes to 1-, 10- and 100- μ g-stimulus load of these five compounds showed that there was a dose effect on the EAG responses for linalool, 3-carene and α -pinene, but not for γ -terpinene and o -xylene. There were no sexual differences between male and female EAG responses to any of the compounds tested. Y-tube olfactometer bioassays of the compounds showed that males and females were attracted to α -pinene, 3-carene, γ -terpinene and/or linalool at doses of 1 and 10  μ g, while weevils were repelled by linalool at a dose of 100  μ g.     

Influence of temperature on silver accumulation and depuration in rainbow trout

To assess the influence of water temperature on silver uptake, rainbow trout Oncorhynchus mykiss ( c . 50 g; held at 13° C) were exposed to 0·1 μM AgNO₃ in ion‐poor water for 1 week at 4 and 16° C without previous temperature acclimation. To assess the influence of temperature on elimination of previously accumulated Ag, rainbow trout were exposed to 0·1 μM AgNO₃ in ion‐poor water for 1 week at 12° C, then were randomly divided amongst two Ag‐free water containers, differing only in temperature (3 and 16° C), for 2 months. In the uptake study greater accumulation of Ag was seen in the gills, plasma and especially the livers and bile of 'warm' rainbow trout (16° C) compared to 'cold' rainbow trout (4° C), which can be explained by the higher metabolic rates of the warmer fish. In the depuration study there was no net elimination of Ag from the livers and bile but there was biphasic elimination of Ag from the gills and plasma of 'warm' and 'cold' fish, but with few differences between them. This indicated that temperature‐dependent processes were less important in Ag elimination than in Ag uptake. Toxicokinetic modelling of Ag uptake by livers indicated four‐fold greater uptake of Ag by 'warm' rainbow trout compared to 'cold' rainbow trout (one compartment uptake model). Elimination of previously accumulated Ag from the plasma was best fitted by a two compartment rate‐constant based model, with approximately half the plasma Ag load eliminated within 24 h, followed by slower elimination of Ag over 2 months.     

Reverse and conventional chemical ecology approaches for the development of oviposition attractants for Culex mosquitoes

Synthetic mosquito oviposition attractants are sorely needed for surveillance and control programs for Culex species, which are major vectors of pathogens causing various human diseases, including filariasis, encephalitis, and West Nile encephalomyelitis. We employed novel and conventional chemical ecology approaches to identify potential attractants, which were demonstrated in field tests to be effective for monitoring populations of Cx. p. quinquefasciatus in human dwellings. Immunohistochemistry studies showed that an odorant-binding protein from this species, CquiOBP1, is expressed in trichoid sensilla on the antennae, including short, sharp-tipped trichoid sensilla type, which house an olfactory receptor neuron sensitive to a previously identified mosquito oviposition pheromone (MOP), 6-acetoxy-5-hexadecanolide. CquiOBP1 exists in monomeric and dimeric forms. Monomeric CquiOBP1 bound MOP in a pH-dependent manner, with a change in secondary structure apparently related to the loss of binding at low pH. The pheromone antipode showed higher affinity than the natural stereoisomer. By using both CquiOBP1 as a molecular target in binding assays and gas chromatography-electroantennographic detection (GC-EAD), we identified nonanal, trimethylamine (TMA), and skatole as test compounds. Extensive field evaluations in Recife, Brazil, a region with high populations of Cx. p. quinquefasciatus, showed that a combination of TMA (0.9 microg/l) and nonanal (0.15 ng/microl) is equivalent in attraction to the currently used infusion-based lure, and superior in that the offensive smell of infusions was eliminated in the newly developed synthetic mixture.     

天牛性信息素、引诱植物和植物性引诱剂的研究与应用

由于目前所拥有的各种天牛防治方法都难以获得理想的防治效果,研发新型天牛控制技术势在必行。其中利用引诱剂是最具潜力的研究领域之一。对天牛的性信息素、引诱植物和植物性引诱剂3个方面的研究及应用作了综述。天牛雌、雄成虫均可释放性信息素进行两性间的通讯联系。天牛性信息素的分泌部位因天牛种类不同而存在着差异。雄、雌成虫触角上均有感受性信息素的感受器,但报道较多的是雄虫感受器。迄今已鉴定出13种天牛性信息素的化学结构。一般而言,天牛成虫的嗜食树种,对天牛具有较强的引诱作用,可作为诱饵树种使用。寄主植物的某些挥发性物质对天牛成虫也有独特的引诱效果。除此之外,对天牛性信息素、引诱植物和植物性引诱剂研究与应用中存在的困难也进行了讨论,并对它们的应用前景做了展望。     

Enhancement of pH‐resistant iron‐binding activity in supernatants of rainbow trout blood leucocytes by in vitro treatment with phorbol‐12‐myristate‐13‐acetate

Leucocyte lysates from rainbow trout Oncorhynchus mykiss showed an iron‐binding activity that was retained even if the samples were exposed to an acid pH (4·5). Iron‐binding activity of leucocyte supernatants was enhanced by the presence of 1 μg ml⁻¹ phorbol‐12‐myristate‐13‐acetate in the cell medium.     

Release of sex steroids into the water by roach

Electro‐olfactogram (EOG) recordings of the olfactory epithelium of both male and female roach Rutilus rutilus demonstrated that both sexes were able to detect free and glucuronidated 17,20β‐dihydroxy‐4‐pregnen‐3‐one (17,20β‐P) with high sensitivity. Male, but not female, roach were also sensitive to androstenedione. Sexually mature female roach were shown to release free 17,20β‐P, glucuronidated 17,20β‐P and androstenedione into the water; for all three steroids, the rate of release was significantly enhanced by injection of carp pituitary extract (CPE). A series of trials was also carried out which showed that mature males, and to a lesser extent immature males and females, were able also to release free and glucuronidated 17,20β‐P, both before and after CPE treatment. Water extracts from containers that had held CPE‐treated mature male and female roach were examined for the presence of other steroids. This revealed that free and glucuronidated 17,20β‐P plus free and glucuronidated 17,20β,21‐trihydroxy‐4‐pregnen‐3‐one (17,20β, 21‐P) predominated in water extracts from both sexes. The free moieties of 17,20α‐dihydroxy‐4‐pregnen‐3‐one, 17‐hydroxyprogesterone and 11‐deoxycortisol were found at concentrations which were between four and 20 times lower than those of free 17,20β‐P. Androstenedione was found at concentrations which were 25‐fold lower than those of 17,20β‐P. Despite its apparent high rate of release by sexually mature male and female roach, free 17,20β,21‐P was found not to exhibit any EOG activity at the highest dose tested (10⁻⁷ M).     

Evaluation of biochemical methods for the non‐destructive identification of sex in upstream migrating salmon and sea trout

Female‐specific markers of reproductive activity [plasma 17β‐oestradiol (E2), vitellogenin (VTG) and alkali‐labile phosphoprotein phosphorous (ALP)] were measured over 12 months in a captive population of brown trout Salmo trutta . During the early months of the reproductive season (February to May) and using the concentration of plasma E2 or plasma ALP as a marker for females the proportion of fish in which sex was misidentified was high (15–50%). The misidentification rate was considerably lower (1–8%) using plasma VTG. Preliminary evaluation of a commercial immunochromatographic VTG test system as a screen for the presence or absence of VTG in plasma from brown trout provided results that were consistent with those obtained from direct measurement of plasma VTG levels by enzyme‐linked immunosorbent assay (ELISA). These preliminary conclusions were verified by sampling upstream‐migrating anadromous brown trout, sea trout, and Atlantic salmon Salmo salar trapped over a 6 month period. Plasma E2 levels did not satisfactorily discriminate between male and female sea trout and Atlantic salmon. Plasma VTG levels in both species, however, were bimodally distributed and it was assumed that this divergence corresponded to male (plasma VTG levels <10 μg ml⁻¹) and female (plasma VTG levels >800 μg ml⁻¹) fishes. Plasma ALP provided a more accurate indication of sex in the wild Atlantic salmon and sea trout than was suggested by the pilot study on captive brown trout. The commercial immunochromatographic VTG test system provided results that were wholly consistent with the data obtained from the trapped fishes by direct measurement of plasma VTG.     

Oviposition by Danaus plexippus in relation to cardenolide content of three Asclepias species in the southeastern U.S.A.

Abstract. 1. Female monarchs were observed in the field ovipositing on a native North American milkweek host, Asclepias humistrata L. As in a comparable Australian study on an introduced novel host ( Asclepias fruticosa L.) we found post-alighting rejection of plants with low and high cardiac glycoside concentration (CG).
2. Most oviposition took place on plants with CG in the range 200–500 μg/0.1 g dry weight. Thin-layer chromatography showed no obvious qualitative difference in cardenolide types between accepted and rejected plants, excepting an indication that rejected plants may have a higher level of more polar cardenolides.
3. In a controlled laboratory experiment comparing oviposition on a low ( A.incarnata L.) vs high ( A.curassavica L.) CG host plant species we found no relationship between CG and oviposition on the low CG species, but a negative relationship in the high CG species. This corroborates our findings on oviposition on single host species in the field.
4. We also record the first indication of a physiological cost of monarch larvae feeding on plants high in CG. There was a significant negative relationship between survival of first instar larvae and CG levels in plants. This study opens the way for further work on the association of monarch butterflies and their toxic milkweed hosts.     

The effect of temperature on the response of Daphnia to toxic cyanobacteria

1. In a set of life‐table experiments, cohorts of neonate Daphnia pulex were exposed to a toxic strain of Anabaena affinis and A. flos ‐ aquae , and to a pure cyanobacterial toxin (anatoxin‐a), at 12 or 14, 19, and 25 °C. The fecundity and survival of individual animals were assessed at 1‐, 2‐ or 3‐day intervals, depending on the temperature, through to the fifth brood of the control cohort. The sensitivity of D . pulex to the cyanobacteria and the toxin at each temperature was measured by determining its finite population growth rate (λ) in an experimental treatment as a fraction of that in a control treatment. Tests with three concentrations of cyanobacteria (1.0, 2.5 and 5.0 μg mL^–1) and one concentration of anatoxin‐a (1 μg mL^–1), and with two clones of D. pulex , showed a consistent and statistically significant pattern of increasing sensitivity at higher temperatures.
2. Anabaena affinis affected both survivorship and fecundity, while A. flos ‐ aquae and its toxin, anatoxin‐a, primarily affected fecundity. Presence of cyanobacteria affected brood size, brood number, time to first reproduction and interclutch interval. Temperature affected time to first reproduction and interclutch interval at all concentrations of cyanobacteria. Brood number and brood size were little affected by temperature except at the highest concentrations of cyanobacteria. Increasing the concentration of A. flos ‐ aquae affected demographic parameters, especially at the lower temperatures, while increasing the concentration of A. affinis had less effect.
3. The study suggests that increasing water temperatures in natural systems should exacerbate the inhibitory effect of toxic cyanobacteria on daphniid population growth rates.     

Olfactory Sex Attraction and Mating Behaviour in the Pine Sawyer Monochamus galloprovincialis (Coleoptera: Cerambycidae)

The pine sawyer Monochamus galloprovincialis Olivier (Coleoptera: Cerambycidae) is a vector of the pine wood nematode Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle. Male and females of this species have a kairomonal attraction to host volatiles and Ips semiochemicals. Once on the host tree, males and females copulate and oviposition occurs. Bioassays using Y-tube olfactometry revealed that females were attracted to volatile compounds produced by males, but not to volatiles produced by females. However, immature males did not seem to release attractants for mature females and immature females did not show any attraction for mature males. A experiment designed to know about mating behaviour in this specie revealed that most of the males encountered females while they were walking. In all pairs both sexes contacted with antennae before copulation, however, in some occasions copulation did not proceed just after antennal contact. On the contrary, in all pairs observed, the male licked the elytra of the female with his mouth palpi just before copulation (licking); immediately afterwards, the male grabbed the female with his forelegs and mounted her suggesting a chemically mediated mate recognition by males.     

Vitellogenin concentrations in the haemolymph and ovaries of Ixodes scapularis ticks during vitellogenesis

The vitellogenin and vitellin concentrations in the haemolymph and ovaries of Ixodes scapularis females were determined using a double sandwich enzyme-linked immunosorbent assay. The level of vitellogenin in the haemolymph began to increase just prior to tick detachment from the host and continued to increase until 2 days after detachment. There was a slight decrease in the vitellogenin level 4 days after detachment, but a second peak was observed approximately 5 days after oviposition. Subsequent to oviposition, the vitellogenin levels in the haemolymph quickly decreased. The concentration of vitellogenin in the haemolymph ranged from 1.55 to 11.48 g l^-1 during the period after dropping from the host through oviposition. The concentration of vitellin in the ovaries began to increase as the female began rapid engorgement (0.03 mg per female) and declined after oviposition (0.1 mg per female).     

Fitness of wild‐caught Drosophila melanogaster females: allozyme variants of GPDH, ADH, PGM, and EST

We have collected several hundred Drosophila melanogaster flies (near Davis, California), isolated them individually, without anesthesia, at the collecting site, and estimated the fitness components of the wild‐caught females under different environmental conditions. The fitness parameters measured are fecundity, oviposition rate, and productivity (egg‐to‐adult viability, development rate, and number of progeny). The environmental variables are two temperatures (22°C and 28°C) and two densities (‘scant’ and ‘crowded’). After the fitness measurements are completed for each individual female, its genotype is determined at four loci encoding enzymes: GPDH and ADH, located on chromosome II and PGM and EST‐C, located on chromosome III. Density has a large significant effect on productivity; temperature has significant effects on fecundity, oviposition rate, and development rate. The experiments show that allozyme polymorphisms are associated with selection effects. Fitness differences between allozyme genotypes occur for all fitness components, except oviposition rate. But which genotype is superior depends on the environmental conditions; heterozygotes exhibit higher fitness than homozygotes in a number of cases, but inferior in others. A unique feature of the present experiments is that the experimental flies are wild‐caught females rather than laboratory‐bred individuals. This revised version was published online in July 2006 with corrections to the Cover Date.     

Orange-derived stimuli regulating oviposition in the Mediterranean fruit fly

Abstract:  Some visual and olfactory host stimuli influencing oviposition of the Mediterranean fruit fly ( Ceratitis capitata Wiedemann) in orange fruit were investigated in a laboratory-reared strain of this species. Mated females of C. capitata were found to be attracted to the same extent by fragrant orange fruits and odourless sham oranges, while unmated females were notably less attracted than mated females by the above objects. Mated females laid significantly more eggs in orange-coloured than in whitish (optically neutral) spherical wax dummies (diameter, 9.0 cm) as well as in both orange-coloured and whitish wax dummies supplemented with high internal humidity (80–100%) compared with respective wax dummies supplied with low internal humidity (30–50%). The orange-like colour together with high internal humidity provides basic stimuli supporting adequate oviposition of C. capitata in oranges. The oviposition rate of female C. capitata was not significantly changed when graded dosages ranging from 0.004 to 19.6  μ l of orange peel oil per cm² were added to orange-coloured wax dummies, while oviposition was considerably subdued by addition of 3.9  μ l orange peel oil and completely disrupted by addition of 9.8 or 19.6  μ l of orange peel oil per cm² of whitish wax dummies. Oviposition of C. capitata in ripe orange fruit may thus be interpreted by the predilection of this tephritid species for an orange-coloured, glossy pericarp, being capable of counteracting the deterrent effect of the essential oil found in orange flavedo.     

Measurements of phosphorus uptake by macrophytes and epiphytes from the LaPlatte River (VT) using 32P in stream microcosms

1. Phosphorus (P) uptake by macrophytes and epiphytes from the LaPlatte River (VT) was examined in the laboratory by adding ³²PO₄‐P to recirculating stream microcosms.
2. Water, plugs of sediment and plants were removed from the river and placed into the microcosms. ³²PO₄‐P was then added either to the water or the sediment, and its incorporation into plants and epiphytes was monitored over 3 days. Uptake was examined at both ambient (5 μg L^–1) and increased (50 μg L^–1) soluble reactive phosphorus (SRP) concentrations. A computer program was developed to fit curves to the radiotracer data and calculate rate constants for the simultaneous transfer of ³²P among compartments.
3. Both macrophytes and epiphytes removed P from the water, but epiphyte uptake of P was more rapid. Phosphate enrichment stimulated P uptake by both macrophytes and epiphytes. Macrophytes also obtained P from the sediment. The relative contribution of P to macrophytes from the water vs. that from the sediment appeared to vary with SRP in the overlying water. Accurate estimates of rates of P uptake from sediments by macrophytes were difficult to obtain however, due to very low and highly variable unit rate constants for P uptake and uncertainty about the magnitude of the phosphate pool available for uptake.
4. SRP concentrations were greater in the overlying water than in the sediment pore water of stream microcosms in the present study. Numerous reports in the literature have suggested that this condition favours uptake by macrophyte stems and leaves rather than by roots.
5. Phosphate uptake from the water by macrophytes in shallow streams may be more common than for macrophytes in lakes.