Synthesis of Mono- and Sesquiterpenoids

The synthesis of isocitrate lyase in Candida tropicalis, the growth of which was stimulated by exogenously added biotin, was released from repression by glucose under biotin-deficient conditions. Biotin deficiency reduced remarkably the levels of biotin-enzymes, pyruvate carboxylase and acetyl-Co A carboxylase, in the glucose-utilizing cells of this yeast. A marked increase in intracellular level of pyruvate was observed in the biotin-deficient cells. Acetyl-CoA-donating compounds, such as pyruvate, acetate and alkanes, stimulated the formation of isocitrate lyase in the yeast regardless of the presence or absence of biotin. On the other hand, malate and succinate did not affect the enzyme synthesis. The isocitrate lyase synthesis under biotin-sufficient conditions was repressed by not only glucose but also glucosamine and 2-deoxyglucose. This repression by glucose was not eliminated by cAMP. The stimulated synthesis of isocitrate lyase under biotin-deficient conditions was also observed in C. albicans and C. guilliermondii growing on glucose.     

Mammalian pyruvate carboxylase: effect of biotin on the synthesis and translocation of apo-enzyme into 3T3-L adipocyte mitochondria

The effect of biotin on the induction (and possible requirement for uptake into mitochondria) of apopyruvate carboxylase has been examined in 3T3-L adipocytes. Cells fed biotin-sufficient medium contained only holoenzyme in mitochondria and no apoenzyme was detected. The amount of apoenzyme elaborated in biotin-deficient 3T3-L adipocytes was comparable to the holopyruvate carboxylase protein found in cells maintained on biotin-sufficient medium. Like the holoenzyme, the apoenzyme was detected exclusively in the mitochondrial fraction of 3T3-L adipocytes. This indicates that the synthesis of apopyruvate carboxylase and its translocation into mitochondria occur independently of the cofactor, biotin.     

Biotin uptake, utilization, and efflux in normal and biotin-deficient rat hepatocytes.

Biotin uptake, utilization, and efflux were studied in normal and biotin-deficient cultured rat hepatocytes. Biotin-deficient cells accumulate about 16-fold more biotin than do normal cells when incubated with a physiological concentration of biotin for 24 h. This difference is due to the greater amount of protein-bound biotin relative to free biotin in biotin-deficient hepatocytes, and is attributable to the presence of more apocarboxylases in deficient cells. The rate of biotin uptake and the rate of activation of the carboxylases, acetyl-CoA carboxylase, pyruvate carboxylase, propionyl-CoA carboxylase, and beta-methylcrotonyl-CoA carboxylase, are proportional to the concentration of exogenous biotin. Increases in carboxylase activities are proportional to the concentration of biotin only at exogenous biotin concentrations of less than 410 nM. Concentrations of 410 nM or more biotin increase carboxylase activities to normal or near normal. Biocytin inhibits biotin uptake at very high concentrations, whereas desthiobiotin and lipoic acid have no effect. Biocytin in the medium results in carboxylase activation either intracellularly or extracellularly by conversion to biotin by biotinidase. Investigation of the efflux of biotin from normal and biotin-deficient cells preincubated with the vitamin showed greater retention of biotin by biotin-deficient cells than by normal cells over 24 h. Retention of free biotin is similar in biotin-deficient and normal cells. The greater amount of biotin retained by biotin-deficient cells is accounted for by the greater amount of bound biotin in these cells. These results suggest that the free and bound biotin pools are independently regulated. The ready loss of free biotin from these cells has implications for the treatment of inherited, biotin-responsive carboxylase deficiencies.     

Drosophila melanogaster larvae fed by glucose and fructose demonstrate difference in oxidative stress markers and antioxidant enzymes of adult flies

Activities of antioxidant and associated enzymes, and oxidative stress markers were assessed in newly enclosed adult fruit flies Drosophila melanogaster developed on diets with 4 and 10% glucose or fructose. In fly males, 10% fructose promoted higher content of protein carbonyls and catalase activity, but lower superoxide dismutase (SOD) activity than 4%, while in females-lower levels of high molecular mass thiols (H-SH). Females at all diets had virtually the same level of lipid peroxides, low-molecular-mass thiols, catalase, and superoxide dismutase activities. Fed with 4% fructose and glucose males demonstrated 24 and 26% lower H-SH level than females, respectively. On diets with 4% glucose, 10% glucose and fructose females had 32, 26 and 27% lower catalase activity than respective males, and 1.3-1.5-fold lower glucose-6-phosphate dehydrogenase activity on glucose-containing diets. Strong positive correlations between H-SH level and G6PD activity, as well as between catalase and G6PDH activity were found. These results suggest that type and concentration of dietary carbohydrate affect antioxidant defense in fruit flies. It also substantially depends on fly sex, comprising presumably levels of protein carbonyls and lipid peroxides, as well as catalase and SOD activities in males and G6PDH activity in females.     

Enzymatic determination of biotin.

An enzymatic method for the quantitative determination of biotin has been developed. The method involves the enzymatic binding of biotin in situ to the pyruvate carboxylase apoprotein of biotin-deficient bakers' yeast and the subsequent estimation of the pyruvate carboxylase activity by a ¹⁴CO₂-fixation method. The method is specific for biotin. Several biotin analogs and precursors were tested, and only biocytin was found to interfere, Biotin amounts of less than 5 pg can be estimated.     

Mitochondria-targeted plastoquinone derivative SkQ1 increases early reproduction of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> at the cost of early survival

It was previously found that 20 pM SkQ1 (10-(6′-plastoquinonyl) decyltriphenylphosphonium) solution increased lifespan of virgin females and males of the fruit fly Drosophila melanogaster with maximal and highly reproducible effect on early survival of females. In this paper we demonstrate that SkQ1 solution of the same concentration does not increase lifespan of mated females and males, early effect on female survival being absent, whereas early fertility and the total number of progeny are elevated in treated flies. Increase in fertility observed in young mated females instead of increase of survival typical for young virgin females might illustrate the trade-off between the fly’s lifespan and reproduction.     

Fecundity,fertility and reproductive recovery of irradiated Queensland fruit fly Bactrocera tryoni

Pupae of the Queensland fruit fly or Q‐fly Bactrocera tryoni (Froggatt) are irradiated routinely to induce reproductive sterility in adults for use in sterile insect technique programmes. Previous studies suggest that adult sexual performance and survival under nutritional and crowding stress are compromised by the current target dose of radiation for sterilization (70–75 Gy), and that improved mating propensity and survival under stress by irradiated males may be achieved by reducing the target sterilization dose without reducing the level of induced sterility. This raises the question of the amount by which the irradiation dose can be reduced before residual fertility becomes unacceptable. The present study measures the levels of residual fertility in male and female irradiated Q‐flies at different irradiation doses (20, 30, 40, 50, 60 and 70 Gy), and investigates the possibility that fecundity and fertility increase between 10–15 and 30–35 days post emergence. Male flies require a higher dose than females to induce sterility, with no residual fertility found in females irradiated at doses of 50 Gy or above, and no residual fertility found in males irradiated at doses of 60 Gy or above. Irradiated females are more fecund at 30–35 days post emergence than at 10–15 days. However, fertility does not increase between 10 and 15 days post emergence and 30–35 days, even at doses below 50 Gy. The present study shows that there is scope to reduce the target sterilization dose for Q‐flies below that of the current dose range (70–75 Gy) at the same time as retaining an adequate safety margin above radiation doses at which residual fertility can be expected.     

Terahertz radiation influence on stressed drosophila life span

Virgin fruit fly females and males were stressed by placement into a confined space without food for 3 h. Part of stressed flies were subjected to terahertz irradiation (0.1–2.2 THz) during 30 min. Life span of individual flies was evaluated. Terahertz radiation had some positive influence on male survival during the stage of monotonic decrease in flies number and negative effect during the stage of relatively stable number of flies. The survival of irradiated females on the stage of sharp decline in the number of flies was higher than in stressed and control females. Authors propose that terahertz radiation has an indirect effect on gene expression and signaling pathways which control the survival and life span of Drosophila.     

Biotin uptake in cultured hepatocytes from normal and biotin-deficient rats

Biotin uptake was studied in isolated cultured hepatocytes of normal and biotin-deficient rats. Biotin uptake was temperature-dependent with respect to physical, but not to chemical, processes, proportional to the exogenous biotin concentration in the medium, independent of pH and sodium ion concentrations of the medium, and uneffected by the presence of structural analogues of biotin or metabolic inhibitors in both normal and biotin-deficient hepatocytes. These results suggest that biotin uptake occurs by a passive, nonmediated, non-energy-dependent mechanism in rat hepatocytes.     

Nectarine promotes longevity in Drosophila melanogaster

Fruits containing high antioxidant capacities and other bioactivities are ideal for promoting longevity and health span. However, few fruits are known to improve the survival and health span in animals, let alone the underlying mechanisms. Here we investigate the effects of nectarine, a globally consumed fruit, on life span and health span in Drosophila melanogaster. Wild-type flies were fed standard, dietary restriction (DR), or high-fat diet supplemented with 0-4% nectarine extract. We measured life span, food intake, locomotor activity, fecundity, gene expression changes, and oxidative damage indicated by the level of 4-hydroxynonenal-protein adduct in these flies. We also measured life span, locomotor activity, and oxidative damage in sod1 mutant flies on the standard diet supplemented with 0-4% nectarine. Supplementation with 4% nectarine extended life span, increased fecundity, and decreased expression of some metabolic genes, including a key gluconeogenesis gene, PEPCK, and oxidative stress-response genes, including peroxiredoxins, in female wild-type flies fed the standard, DR, or high-fat diet. Nectarine reduced oxidative damage in wild-type females fed the high-fat diet. Moreover, nectarine improved the survival of and reduced oxidative damage in female sod1 mutant flies. Together, these findings suggest that nectarine promotes longevity and health span partly by modulating glucose metabolism and reducing oxidative damage.     

Targeted expression of the human uncoupling protein 2 (hUCP2) to adult neurons extends life span in the fly

The oxidative stress hypothesis of aging predicts that a reduction in the generation of mitochondrial reactive oxygen species (ROS) will decrease oxidative damage and extend life span. Increasing mitochondrial proton leak-dependent state 4 respiration by increasing mitochondrial uncoupling is an intervention postulated to decrease mitochondrial ROS production. When human UCP2 (hUCP2) is targeted to the mitochondria of adult fly neurons, we find an increase in state 4 respiration, a decrease in ROS production, a decrease in oxidative damage, heightened resistance to the free radical generator paraquat, and an extension in life span without compromising fertility or physical activity. Our results demonstrate that neuronal-specific expression of hUCP2 in adult flies decreases cellular oxidative damage and is sufficient to extend life span.     

四种不同来源的超氧化物歧化酶（SOD）对黑腹果蝇寿命、 繁殖力和抗逆能力的影响

【目的】 评价不同来源的超氧化物歧化酶（SOD）对果蝇延长寿命, 增强繁殖力和抗逆能力的功效。【方法】以黑腹果蝇Drosophila melanogaster为实验材料, 比较研究从家蝇Musca domestica中提取的SOD、 在毕赤酵母Pichia pastoris中重组表达的人hEC SOD、 在酿酒酵母Saccharomyces cerevisiae中重组表达的中国拟青霉Paecilomyces sinensis SOD (ps-SOD)以及商品SOD对果蝇寿命、 繁殖力和抗逆能力的影响。【结果】在饲料中添加4种SOD均能显著延长果蝇的平均寿命, 雌果蝇寿命延长8.09%~12.38%, 雄果蝇寿命延长12.01%~15.86%; F1代雌性子代数量增加25.94%~30.07%, 雄性增加21.75%~39.54%。果蝇的耐高温和抗紫外辐射能力与添加的SOD浓度有关。在饲料中添加较高剂量的SOD, 使热暴露雌性果蝇的寿命延长7.45%~9.88%, 雄性果蝇延长13.46%~15.12%; 受紫外线辐射的雌性果蝇的寿命延长13.47%~20.47%, 雄性果蝇延长16.49%~23.73%。【结论】综合评价认为, 4种SOD均能延长果蝇寿命, 增强其繁殖力和抗逆能力, 但这些功效在本研究供试的4种SOD间无显著差异, 为不同来源SOD的应用提供了重要数据。     

Effects of larval diet on development rates and reproductive maturation of male and female Mediterranean fruit flies

Abstract Mediterranean fruit fly Ceratitis capitata (Diptera: Tephritidae) larvae are capable of developing in one of many hosts that may vary greatly in quality. We hypothesized that they will respond to the larval environment in a manner beneficial to their subsequent reproductive performance. Accordingly, we investigated the effects of various larval diets (varying in the amount of protein and sugar they contain) on the size, development time, nutritional status and reproductive maturation (ovarian development and onset of sexual behaviour) of females and males. We found that flies which undergo larval development in artificial host fruit that contain sugar and protein ('protein-fed') were larger, developed faster and emerged with more nutritional reserves than flies that were protein-deprived as larvae. Protein-fed males, regardless of their size, became sexually active before males that developed in hosts with no protein. Protein-fed females produced more mature eggs than protein-deprived ones. Moreover, protein-fed females tended to copulate sooner than females that developed in hosts with no protein. In addition, regardless of female larval diet, females with more mature eggs tended to copulate sooner than females with less mature eggs. In light of these results, the importance of the larval environment for adult reproductive success is discussed.     

Seasonal morphotypes of Drosophila suzukii differ in key life‐history traits during and after a prolonged period of cold exposure

Seasonal polyphenism in Drosophila suzukii manifests itself in two discrete adult morphotypes, the “winter morph” (WM) and the “summer morph” (SM). These morphotypes are known to differ in thermal stress tolerance, and they co‐occur during parts of the year. In this study, we aimed to estimate morph‐specific survival and fecundity in laboratory settings simulating field conditions. We specifically analyzed how WM and SM D. suzukii differed in mortality and reproduction during and after a period of cold exposure resembling winter and spring conditions in temperate climates. The median lifespan of D. suzukii varied around 5 months for the WM flies and around 7 months for the SM flies. WM flies showed higher survival during the cold‐exposure period compared with SM flies, and especially SM males suffered high mortality under these conditions. In contrast, SM flies had lower mortality rates than WM flies under spring‐like conditions. Intriguingly, reproductive status (virgin or mated) did not impact the fly survival, either during the cold exposure or during spring‐like conditions. Even though the reproductive potential of WM flies was greatly reduced compared with SM flies, both WM and SM females that had mated before the cold exposure were able to continuously produce viable offspring for 5 months under spring‐like conditions. Finally, the fertility of the overwintered WM males was almost zero, while the surviving SM males did not suffer reduced fertility. Combined with other studies on D. suzukii monitoring and overwintering behavior, these results suggest that overwintered flies of both morphotypes could live long enough to infest the first commercial crops of the season. The high mortality of SM males and the low fertility of WM males after prolonged cold exposure also highlight the necessity for females to store sperm over winter to be able to start reproducing early in the following spring.     

Overexpression of glutamate-cysteine ligase extends life span in Drosophila melanogaster

The hypothesis that overexpression of glutamate-cysteine ligase (GCL), which catalyzes the rate-limiting reaction in de novo glutathione biosynthesis, could extend life span was tested in the fruit fly, Drosophila melanogaster. The GAL4-UAS binary transgenic system was used to generate flies overexpressing either the catalytic (GCLc) or modulatory (GCLm) subunit of this enzyme, in a global or neuronally targeted pattern. The GCL protein content of the central nervous system was elevated dramatically in the presence of either global or neuronal drivers. GCL activity was increased in the whole body or in heads, respectively, of GCLc transgenic flies containing global or neuronal drivers. The glutathione content of fly homogenates was increased by overexpression of GCLc or GCLm, particularly in flies overexpressing either subunit globally, or in the heads of GCLc flies possessing neuronal drivers. Neuronal overexpression of GCLc in a long-lived background extended mean and maximum life spans up to 50%, without affecting the rate of oxygen consumption by the flies. In contrast, global overexpression of GCLm extended the mean life span only up to 24%. These results demonstrate that enhancement of the glutathione biosynthetic capability, particularly in neuronal tissues, can extend the life span of flies, and thus support the oxidative stress hypothesis of aging.     

Early Life Hormetic Treatments Decrease Irradiation-Induced Oxidative Damage,Increase Longevity,and Enhance Sexual Performance during Old Age in the Caribbean Fruit Fly

Early life events can have dramatic consequences on performance later in life. Exposure to stressors at a young age affects development, the rate of aging, risk of disease, and overall lifespan. In spite of this, mild stress exposure early in life can have beneficial effects on performance later in life. These positive effects of mild stress are referred to as physiological conditioning hormesis. In our current study we used anoxia conditioning hormesis as a pretreatment to reduce oxidative stress and improve organismal performance, lifespan, and healthspan of Caribbean fruit flies. We used gamma irradiation to induce mild oxidative damage in a low-dose experiment, and massive oxidative damage in a separate high-dose experiment, in pharate adult fruit flies just prior to adult emergence. Irradiation-induced oxidative stress leads to reduced adult emergence, flight ability, mating performance, and lifespan. We used a hormetic approach, one hour of exposure to anoxia plus irradiation in anoxia, to lower post-irradiation oxidative damage. We have previously shown that this anoxic-conditioning treatment elevates total antioxidant capacity and lowers post-irradiation oxidative damage to lipids and proteins. In this study, conditioned flies had lower mortality rates and longer lifespan compared to those irradiated without hormetic conditioning. As a metric of healthspan, we tracked mating both at a young age (10 d) and old age (30 d). We found that anoxia-conditioned male flies were more competitive at young ages when compared to unconditioned irradiation stressed male flies, and that the positive effects of anoxic conditioning hormesis on mating success were even more pronounced in older males. Our data shows that physiological conditioning hormesis at a young age, not only improves immediate metrics of organismal performance (emergence, flight, mating), but the beneficial effects also carry into old age by reducing late life oxidative damage and improving lifespan and healthspan.     

Prevalence of Candidatus Erwinia dacicola in wild and laboratory olive fruit fly populations and across developmental stages

The microbiome of the olive fruit fly, Bactrocera oleae (Gmelin), a worldwide pest of olives (Olea europaea L.), has been examined for >100 yr as part of efforts to identify bacteria that are plant pathogens vectored by the fly or are beneficial endosymbionts essential for the fly's survival and thus targets for possible biological control. Because tephritid fruit flies feed on free-living bacteria in their environment, distinguishing between the transient, acquired bacteria of their diet and persistent, resident bacteria that are vertically transmitted endosymbionts is difficult. Several culture-dependent and -independent studies have identified a diversity of species in the olive fruit fly microbiome, but they have not distinguished the roles of the microbes. Candidatus Erwinia dacicola, has been proposed to be a coevolved endosymbiont of the olive fruit fly; however, this was based on limited samples from two Italian populations. Our study shows that C. Erwinia dacicola was present in all New and Old World populations and in the majority of individuals of all life stages sampled in 2 yr. Olive fruit flies reared on olives in the laboratory had frequencies of C. Erwinia dacicola similar to that of wild populations; however, flies reared on artificial diets containing antibiotics in the laboratory rarely had the endosymbiont. The relative abundance of C. Erwinia dacicola varied across development stages, being most abundant in ovipositing females and larvae. This uniform presence of C. Erwini dacicola suggests that it is a persistent, resident endosymbiont of the olive fruit fly.     

Oxidative stress adaptation with acute,chronic, and repeated stress

Oxidative stress adaptation, or hormesis, is an important mechanism by which cells and organisms respond to, and cope with, environmental and physiological shifts in the level of oxidative stress. Most studies of oxidative stress adaption have been limited to adaptation induced by acute stress. In contrast, many if not most environmental and physiological stresses are either repeated or chronic. In this study we find that both cultured mammalian cells and the fruit fly Drosophila melanogaster are capable of adapting to chronic or repeated stress by upregulating protective systems, such as their proteasomal proteolytic capacity to remove oxidized proteins. Repeated stress adaptation resulted in significant extension of adaptive responses. Repeated stresses must occur at sufficiently long intervals, however (12-h or more for MEF cells and 7 days or more for flies), for adaptation to be successful, and the levels of both repeated and chronic stress must be lower than is optimal for adaptation to acute stress. Regrettably, regimens of adaptation to both repeated and chronic stress that were successful for short-term survival in Drosophila nevertheless also caused significant reductions in life span for the flies. Thus, although both repeated and chronic stress can be tolerated, they may result in a shorter life.     

Sex differences in movement between natural feeding and mating sites and tradeoffs between food consumption,mating success and predator evasion in Mediterranean fruit flies (Diptera: Tephritidae)

Summary Systematic quantitative observations of the location and diel pattern of adult Mediterranean fruit fly, Ceratitis capitata (Wiedemann), activities were carried out in an orange grove and surroundings on the island of Chios in Greece. Natural fly foods were assessed for their contribution to fly longevity, fecundity and fertility. There were diel shifts in male and female location. Females required a substantial and varied diet to realize peak fecundity. This diet was acquired away from the primary host, orange. Foraging for food throughout most of the day on fig and non-host foliage (including feeding on bird droppings) as well as on fig fruit and grapes, females dispersed and fed more than males. A diet of grapes alone did not support any fecundity, contributing only to longevity. A diet of figs alone, on the other hand, sustained both longevity and egg production. Bird feces alone supported neither egg production nor longevity. However, when added to a diet of figs, bird feces significantly increased fly fecundity. Throughout most of the day, males aggregated in leks within the inner canopy of the primary host, orange. The arrival here during the warmest hours of the day of receptive females, followed by pair formation, reinforced the lek mating system on host foliage. In the afternoon, females shifted to orange fruit where they suffered from high predation mortality while ovipositing. Soon after, males also shifted to orange fruit, where they attempted matings with non-receptive ovipositing females. Male feeding on fig fruit occurred late in the day, a time when they were least likely to find a mate. Male survival did not differ between the natural diets. Tradeoffs between food consumption, mating success and predator evasion are discussed for each sex and related to fruit fly mating systems.