Entomopathogenic activities of environmental isolates of Bacillus thuringiensis against dipteran larvae.

Twenty-five field isolates of Bacillus thuringiensis from the Lower Silesia region of Poland, the Osola plain, and phylloplane niches and soil samples from the Karkonosze National Park were tested for insecticidal activity against larvae of the dipterans Aedes aegypti, Drosophila melanogaster and Musca domestica. The spore-crystal mixture of B. thuringiensis finitimus (soil isolate) killed nearly half of the fourth instar larvae of Ae. aegypti. Similar mortality of between 39-61% was recorded after treatment with two phylloplane isolates belonging to biochemical type I of B. thuringiensis (subsp. japonensis, yoso and jinghongiensis) and type IV (subsp. tochigiensis,). The effect of B. thuringiensis wratislaviensis H-47 was lowest, with larval mortality ranging from 7-28%. The susceptibility of D. melanogaster, evaluated as percentage of larvae that did not survive to adulthood, varied between 0-38% and 0-45% at dosages of 0.5 and 1 ml, respectively. The LC50s ranged from 5.07 x 10(9) to 1.04 x 10(11) spore-crystal mixture. The LC50s for M. domestica larvae were 4.1-4.8 x 10(8) spores/ml of B. thuringiensis finitimus and the phylloplane strain KpC1 (I type) and 2.9 x 10(9) for the phyloplane representative of group I. Generally, all tested isolates showed the most activity against common fly larvae. The highest (64-81% mortality) was observed after treatment with both phylloplane (OpPs1, KpF3, KpC1) and soil isolates (KsAc1) of biochemical type I as well as B. thuringiensis finitimus (type III).

Sexual receptivity in Anopheles gambiae mosquitoes: absence of control by male accessory gland substances.

Although male accessory gland substances modulate female receptivity in most mosquito genera that have been studied, they are not active in the development of sexual refractoriness in the malaria vectors, Anopheles gambiae or Anopheles albimanus. Neither the implantation of male accessory glands nor the injection of gland homogenates affected the insemination rate of unmated females. Similarly, portions of the male genital tract and spermathecae from mated females were also inactive when introduced into these females. Interruption of nervous transmission from the brain reduced the tendency of females to become inseminated and removal of the terminal abdominal ganglion completely abolished insemination. Oviposition behavior did not occur unless the spermatheca containing sperm was present, suggesting that this may be the mechanism that mediates the switchover to mated behavior.

Age of Anopheles gambiae Giles male mosquitoes at time of mating influences female oviposition.

Female Anopheles gambiae were more likely to oviposit when they mated with males that were two days old than when they mated with older males. The females' age at the time of mating and blood feeding did not influence ovipositional patterns. Females were no less likely to oviposit when they mated after blood feeding and maturing eggs. Sperm were present in the seminal vesicles of males 28 hours after eclosion, and their accessory glands were replete by 76-100 hours. The mean life span of sucrose-fed unmated females and males was 22 days.

The check is in the male: male mosquitoes affect female physiology and behavior.

The accessory glands of male mosquitoes may produce substances that are transferred to females during mating and alter female physiology and behavior. The effects of male substances include the inhibition of subsequent female mating behavior, stimulation of oviposition and preoviposition behaviors, and the inhibition of host-seeking behavior. The circadian rhythmicity of females can also be altered and their metabolic priorities restructured, making them more likely to reproduce. The specific components that affect the female have yet to be completely identified, but the published reports are summarized.

A male accessory gland protein that modulates female mosquito (Diptera: Culicidae) host-seeking behavior.

The male accessory gland product that modulates the host-seeking behavior of female Aedes aegypti (L.) mosquitoes was found to be a peptide of 7,600 mw. This peptide also prevented subsequent mating behavior and weakly stimulated oviposition. Neither whole glands nor gland fractions from Anopheles gambiae had any effect on Ae. aegypti females, but those from Aedes albopictus were active.

Effect of body size on host seeking and blood meal utilization in Anopheles gambiae sensu stricto (Diptera: Culicidae): the disadvantage of being small.

The survival, metabolic reserves, and host-seeking response of unfed Anopheles gambiae Giles sensu stricto females of different size classes were investigated from 1 to 6 d after emergence. In addition, the effect of blood meal size and frequency of feeding on the accumulation of metabolic reserves and reproductive development were investigated during the 1st gonotrophic cycle. Large females survived longer and contained significantly more protein, glycogen, and lipid at emergence than did small females. Over 6 d, the host-seeking response of large mosquitoes was greater than that of small ones. There was no gonotrophic development in unfed small females after emergence, whereas < or = 52% of large females developed oocytes to Christophers stage II. One blood meal, given by enema, resulted in gonotrophic development to stage II in small females, whereas only large females matured oocytes to Christophers stage V. In both large and small females, egg development was related to the size of the blood meal, but large females were better able to utilize the meals. Two successive blood meals enhanced the ability of small females to develop eggs. The pregravid condition commonly observed in An. gambiae appears to be determined by insufficient metabolic reserves at emergence, and the probability for small individuals to be pregravid is greater than for large mosquitoes because of the need to build up sufficient energy reserves before gonotrophic development can occur. The consequence of this behavior in relation to malaria transmission is discussed.

Reduced transfer of male accessory gland proteins and monandry in female Aedes aegypti mosquitoes.

We examined whether female Aedes aegypti (L.) mosquitoes would remate after they first mated with a male that was reared on a suboptimal larval diet and that as a result, transferred reduced amounts of male accessory gland proteins. Accessory gland proteins from males labeled with 3H leucine were not detected in females allowed to male mate with the labeled males after they first mated with either low diet or high diet males. The amount of the male accessory gland protein transferred by smaller, low diet males was adequate to terminate female receptivity, even after one gonotrophic cycle, and females of this species appear to be monogamous.

Methoprene affects the rotation of the male terminalia of Aedes aegypti mosquitoes.

The rotation of the male terminalia of Aedes aegypti mosquitoes was investigated with chemical surgical, and mechanical methods. Neither decapitation nor ventral nerve cord transection affected the postemergence rotation, but methoprene administered to 4th-instar larvae and early instar pupae slowed rotation significantly and, in the case of pupal treatment, also interfered with successful insemination. Methoprene appeared to interfere with the developmental maturation of the cross muscles located between the 7th and 8th abdominal segment and postemergence nervous or humoral signals did not appear to affect this rotation.

Distention and sugar feeding induce autogenous egg development by the Asian tiger mosquito (Diptera:Culicidae).

Mechanisms initiating autogenous egg development were studied using a selected strain of Asian tiger mosquito, Aedes albopictus (Skuse), that required a sugar meal to develop eggs autogenously. Caloric intake and the abdominal distention produced by ingesting sucrose solutions were interrelated in their effects on autogeny. Distention of the abdomen with 2 microliters of saline, with no caloric intake, induced autogenous egg maturation in 66% of the females. Abdominal distention produced by 2 microliters of saline did not induce egg development if the ventral nerve cord was transected. However, eggs were produced when females ingested 200 micrograms of sucrose in 2 microliters of water following ventral nerve cord transection. A meal containing at least 100 micrograms of sucrose was required for egg development if abdominal distention was < 1 microliter. Mating influenced autogeny in only 10% of the population. Neither distention, caloric intake nor mating affected the number of eggs that matured.

Endogenous factors regulating mosquito host-seeking behaviour.

The physiological state of the mosquito can modulate behaviours that are normally activated by external stimuli. Even though host stimuli may be present at certain times, the insect may not always express host-seeking behaviour, depending upon the physiological factors that predominate. Traditional views of the gonotrophic cycle characterize mosquitoes as engaging in blood feeding only once before depositing eggs. However, physiological state, including such factors as age, nutritional state, presence of eggs, mating condition, circadian rhythmicity and the number of gonotrophic cycles completed, can affect the expression of this behaviour.

Nutritional reserves of autogenous and anautogenous selected strains of Aedes albopictus (Diptera: Culicidae).

Autogeny in Aedes albopictus (Skuse) females was increased from 5% in wild-type parental stock to 84% by selection through 20 generations and was decreased to 0% by selection through nine generations. Concomitantly, the mean number of eggs per female that matured autogenously increased from 6.4 to 18.3 in sugar-fed mosquitoes that developed eggs. Although the duration of the larval period of the two strains was not different, autogenous females emerged with greater amounts of metabolizable protein and lipid and total available energy reserves than did anautogenous females, but with lesser amounts of glycogen. Dry weights did not differ significantly. The difference in the energy reserves between strains was slightly more than the total energy invested in 20 autogenous eggs. Autogenous females survived 1.3 d longer than did anautogenous females when starved from emergence.

Mosquito gonotrophic cycle and multiple feeding potential: contrasts between Anopheles and Aedes (Diptera: Culicidae).

Two aedine and three anopheline species were allowed to blood-feed to repletion, and their host-seeking behavior was measured with an olfactometer at intervals afterward. Both Aedes aegypti (L.) and Ae. albopictus (Skuse) were inhibited from subsequently seeking a host during the gonotrophic cycle, but Anopheles gambiae Giles, An. albimanus Wiedemann, and An. freeborni Aitken, all showed varying degrees of host-seeking behavior while their eggs matured. Continued host-seeking by some anophelines during the gonotrophic cycle has important implications for some assumptions commonly made about mosquito behavior and parasite transmission.

Endogenous regulation of the attraction of Aedes aegypti mosquitoes.

Even when host attractants are present, there are times when endogenous physiological mechanisms prevent female mosquitoes from responding to them and engaging in host-seeking behavior. These times include a brief postemergence period, periods of the day determined by a circadian clock, and following a blood meal before and after eggs develop, which are controlled by nervous and humoral mechanisms. Other factors such as age, nutrition, and mating status can further modulate host-seeking behavior. The interplay of these factors affects the frequency at which mosquitoes will approach a host and feed on its blood, thus affecting the rates of parasite acquisition and transmission.

Mating and nutritional state affect the reproduction of Aedes albopictus mosquitoes.

Mated female Aedes albopictus mosquitoes that were maintained after emergence without carbohydrate were more likely to develop eggs after a small blood meal than were unmated females maintained on the same diet. The effect was due to male accessory gland substances transferred to the female during mating. Neither the endogenous reserves of protein and lipid nor the number of eggs developed per volume of ingested blood differed between mated and unmated females, suggesting that the utilization of existing reserves was altered by mating. Methoprene administered to both mated and unmated females that ingested small blood meals reduced the likelihood that egg development would occur. Small volumes of blood were more likely to trigger oogenesis in both mated and unmated females if their abdomens were additionally distended.

Sublethal effects of larval methoprene exposure on adult mosquito longevity.

Larvae of Aedes aegypti were exposed to sublethal concentrations of the insect growth regulator, methoprene, and the glycogen content of pupae and surviving adults was compared and effects on adult longevity determined. The glycogen reserves in both male and female Ae. aegypti pupae were significantly reduced as a result of methoprene exposure. The longevity of adult females was also significantly reduced, but exposure affected neither the longevity nor the glycogen content of adult males. Adult sugar feeding increased the amount of glycogen in both treated and control females. The reduced longevity of adult females from larval methoprene treatment appeared not to be directly related to reduced glycogen, but rather reflected neuroendocrine abnormalities induced by this juvenile hormone analogue.

Reproductive and metabolic differences between Aedes aegypti and Ae. albopictus (Diptera: Culicidae).

Aedes albopictus (Skuse) females were more likely to develop eggs after they ingested small blood meals than were Aedes aegypti (L.) when both species were maintained under the same suboptimal adult nutritional regimen. The longevity of adult female Ae. albopictus under conditions of starvation was also significantly greater than that of Ae. aegypti. Analyses of total body proteins, lipids, and glycogen indicated that the increased reproductive efficiency of Ae. albopictus may be a result of its greater reserves accumulated during a longer larval period.

A bioassay for cobra cardiotoxin activity using semi-isolated cockroach heart.

A semi-isolated cockroach heart preparation was used to rapidly determine the activity of cobra cardiotoxin, monitored as a direct response on heart rate. This preparation produced a dose-response curve in the presence of active cardiotoxin and demonstrated that cardiotoxin retained its biological activity after boiling, although cardiotoxin activity was destroyed by heating in the presence of dithiothreitol. Experiments that cross-linked radiolabeled cardiotoxin to solubilized cockroach heart membranes suggested that cardiotoxin bound specifically to a 59,000 mol. wt membrane protein in this tissue.

Correlation of nutritional reserves with a critical weight for pupation in larval Aedes aegypti mosquitoes.

The relationships between the readiness of fourth instar Aedes aegypti to metamorphose and their weight, nutritional reserves and sex were examined. Both the critical larval weight required for metamorphosis and the potential weight were influenced by the temperature at which the larvae were reared, and were greater for females than for males. Caloric levels of soluble carbohydrates and soluble lipids correlated strongly with weight. The accumulations of both carbohydrates and lipids were sex-dependent and were increased in both sexes at the higher temperature, but caloric levels of glycogen at the critical weights were all similar. We suggest that weight per se is not a critical factor in determining readiness to pupate, and that carbohydrate levels may be one important causal variable.

The endogenous regulation of mosquito reproductive behavior.

Most female mosquitoes require a meal of blood that provides protein for egg maturation. For reproduction to occur, two behavioral sequences are essential. One is concerned with finding a host for the blood meal and the other in finding a site on which to lay the eggs that result. Stimuli from both hosts and oviposition sites initiate the reproductive behaviors of host-seeking and pre-oviposition, respectively, that are discussed in this review. After sensory receptors perceive these stimuli, the central nervous system must integrate the information and associate it with a biologically appropriate response. Host-seeking appears to be the default behavior, expressed whenever host stimuli are present. However, if the female is successful in locating a host and ingesting blood, subsequent host-seeking is inhibited when the meal distends the abdomen above a certain threshold. Host-seeking inhibition continues during egg development as a result of a humoral mechanism even after the blood volume has been reduced by digestion. At the time when eggs are maturing and host-seeking is inhibited, pre-oviposition behavior predominates if the central nervous system receives oviposition site stimuli. This behavior is also initiated by a humoral factor. Several physiological states, including insemination, age, and nutrition, can modulate both host-seeking and pre-oviposition behaviors.