Further observations on rhythmic emission of fragrance in flowers

Observations regarding floral fragrance and the rhythmicity of its emission in four plant species are reported. In the case of flowers of Hoya carnosa R. Br. which are characterized by circadian rhythmicity of scentedness (R. Altenburger and P. Matile, 1988, Planta 174, 248–252), temperature compensation of the free-running period as well as persistence of oscillations in permanent darkness have been demonstrated. A hitherto unidentified component of fragrance turned out to be identical to an unusual sesquiterpene recently discovered in cardamom oil (B. Maurer et al., 1986, Tetrahedron Lett. 27, 2111–2112). In Stephanotis floribunda Brongs. the rhythmic emission of fragrance is circadian in nature, but in a constant environment the oscillations of individual components are increasingly asynchronous. In excised flowers of Odontoglossum constrictum Lindl. the diurnal oscillations observed in a natural photoperiod are abolished under constant environmental conditions. They are resumed upon the return to a 1212-h photoperiod. The absence of circadian control could also be demonstrated in excised flowers of Citrus medica L. In this species, too, the daily maxima of scent emission reappear upon the transfer of flowers to a 1212-h light/ dark cycle. Results obtained upon the comparative analysis of volatiles in the headspace above the flowers and in petal extract indicate that the relative abundance of an individual compound in the floral fragrance is not a function of differential volatility.Abbreviations DL 1212 h photoperiod - DD continuous darkness - GLC gas-liquid chromatography - LL continuous illumination The authors are indebted to R. Kaiser, Givaudon Corp.for kindly carrying out the identifikation of volatiles by gas-liquid chromalography/mass spectrometry.     

Circadian rhythmicity of fragrance emission in flowers of Hoya carnosa R. Br.

In flowers of Hoya carnosa R. Br. the nocturnal emission of fragrance occurs according to an endogenous circadian rhythmicity. This was demonstrated by the continuation of rhythmical emission for two to three cycles under conditions of permanent illumination. The free-running period was approx. 29 h under these conditions. The inversion of light/dark cycles caused the entrainment of the next peak of emission by approx. 12 h whether the inversion was started with a light period or a with dark period prolonged for 12 h. When only the flower used for fragrance analysis was subjected to an inverted photoperiod, the rest of the plant remaining under the original light/dark cycle, the synchronization of fragrance emission occurred according to the Zeitgeber perceived by the individual flower.     

Circadian rhythmicity in emission of volatile compounds by flowers of Rosa hybrida L. cv. Honesty

Quantitative headspace analyses of rose flowers showed no significant differences in composition of emitted volatile compounds between flowers on the intact plant and cut flowers placed in vase water containing sucrose at 0, 15 or 30 g · l⁻¹. Volatile components emitted were geraniol, nerol, citronellol, E-citral, Z-citral, methylgeranylate, trans-caryophyllene, β-cubebene, dihydro-β-ionone, 2-phenylethanol, 2-phenylethylacetate, 3,5-dimethoxytoluene and hexylacetate. When exposed to a 12-h photoperiod these components showed maximum emission during the light period and a rhythmicity which differed for the individual compounds. The circadian nature of the rhythmicity was confirmed by the continuation of rhythmicity during continued darkness or light, and was characterized by `transient' and `free running' periods of 27 and 24 h, respectively, and a phase shift of 12 h in rhythmicity when a 24-h period of continuous darkness was followed by re-exposure to a 12-h photoperiod. Rhythmicity in emission was not observed when flowers were kept in darkness before flower bud opening, but started immediately upon exposure to a 12-h photoperiod. Received: 14 April 1998 / Accepted: 28 May 1998     

Rhythms of fragrance emission in flowers

A method for the sampling of volatiles emitted by individual flowers is described. Sampling over periods of 3 h allowed the examination of diurnal changes in quantity and quality of fragrance. In the species studied, Odontoglossum constrictum Lindl., Citrus medica L., Hoya carnosa R. Br., and Stephanotis floribunda Brongs., the fragrance was characterized by a few major components accompanied by a larger number of minor components. Flowers of all species produced volatiles in a rhythmical, diurnal fashion. Whereas in detached flowers of O. constrictum and C. medica rhythmicity could be observed for up to four cycles, flowers of H. carnosa showed this phenomenon only when attached to the plant. Maxima of emission were observed during the day in C. medica and O. constrictum whereas in H. carnosa it occurred during the night. In S. floribunda a conspicuous asynchronism of the emission of different volatiles was observed, resulting in the rhythmical change of fragrance quality.     

Ahemeral light regimens test the photoperiodic threshold of the european starling(Sturnus vulgaris)

Male European starlings(Sturnus vulgaris) were held for three consecutive photoperiod oscillations (ahemeral years) composed of 30-h day lengths, i.e., the daily light and dark each lasted three hours longer than under the natural daily photoperiod at latitude 38°N. These starlings had no gonad metamorphosis during the 45 actual months necessary to complete the three ahemeral photoperiod oscillations; nor did subsequent exposure to continuous illumination elicit gonad response. It is concluded that the daily duration of light and darkness (although certainly operant in controlling starling sexual cycles under temperate-zone photoperiod oscillations) is not the critical factor establishing a sexual cycle under the ahemeral regimen. Rather, it appears that this species must experience a daily duration of light of 12 hours or less (a definitive photoperiodic threshold) before photo-induction of a sexual cycle is possible.Presented at the Eighth International Congress of Biometeorology, 9–14 September 1979, Shefayim, Israel.     

CAM-idling in Hoya carnosa (Asclepiadaceae)

In the leaf succulent Asclepiad Hoya carnosa (L.) R. Br., CAM photosynthesis occurred under well-watered conditions, as characterized by diurnal gas exchange and changes in titratable acidity. Following 10–12 days of severe water stress, the plants shifted from CAM to a modified CAM-idling mode of metabolism. CAM-idling was characterized by complete or almost complete stomatal closure accompanied by CAM-like diurnal changes in titratable acidity. H. carnosa plants maintained this CAM-idling mode of photosynthesis for at least 8 weeks. Upon reirrigation, the plants returned to the original CAM mode within 1 week. These results suggested that CAM-idling is a reversible, intermediate form of sustained metabolism which enables plant survival under conditions of extended drought.This work was supported in part by NSF Grant PCM 8200366 and in part by the Science and Education Administration of the United States Department of Agriculture under Competitive Grant 5901-0420-8-0018-0.     

Gene expression profile of Ralstonia Solanacearum for the rhizosphere ecological niche of Solanum tuberosum

Plant root secretion can be regarded as signal molecules, which exerts impact on microorganisms in the rhizosphere ecological niche. We obtained gene expression profile of Ralstonia solanacearumPO41 under the root secretions environment of Solanum tuberosum at the time points of 8 hrs, 16 hrs and 24 hrs, respectively, after infection with RNA microarray technology. Bioinformatics tools of differential genes expression analysis, GO functional analysis, cluster analysis and pathway analysis were conducted to find out the pathogenic genes and other related genes. We found that the virulence factors of R. solanacearum mainly focused on the output pathways of toxic protein (Sec pathway, Tat pathway and type III secretion system (T3SS)), the aggregation and transfer of exopolysaccharides and the chemotactic movement and adhesion of flagellum in the potato root secretion ecological niche, while the virulence factors in the atypical output pathway mainly distributed in Sec (secB, secDF, yidc) and Tat (tatA, tatC) pathways to promote the output of folded and unfolded toxic proteins. The fliIATPase was obviously upregulated 8 hrs postinoculation, suggesting that type III secretion system was only active at the early stage of PO41 infection. The upregulated expression of phosphoglucomutase and epimerase showed that the virulence factor of exopolysaccharides (EPS) was synthesized at the early stage of R. solanacearum infection. Chemotactic receptor and motor protein were obviously upregulated within 24 hrs postinoculation. Our study revealed that R. solanacearumPO41 had already colonized to the roots within 24 hrs with the stimulating of root secretion. Some pathogenic genes were upregulated during this period.     

Effect of photoperiod manipulation on the growth performance of juvenile lenok,Brachymystax lenok (Pallas, 1773)

The effect of four different light regimes on growth was studied in lenok, Brachymystax lenok. Fish with average weights of 5.5 g were subjected to four different photoperiods (0L:24D, 6L:18D, 12L:12D and 24L:0D) for 35 days. The specific growth rate (SGR) of lenok in 24‐h darkness had a significantly higher SGR than those in the continuous light regime (P < 0.05); however, there was no significant difference among fish exposed to 6L:18D, 12L:12D and 24L:0D photoperiods. There was a tendency for higher food intake over the light period extension from 0L to 24L, and feed intake was significantly higher in the continuous light group than in 24‐h darkness (P < 0.05). No significant difference in feed conversion efficiency (FCE) was observed between fish exposed to 0L:24D and 6L:18D photoperiods, however, the FCE in both photoperiods was significantly higher than that in the other two groups. The final survival rate of juveniles varied from 79.67 to 95.33%, with significant differences among experimental groups. Fish tested in continuous illumination spent much more energy on respiration and excretion while depositing less energy for growth than in the other photoperiods. In contrast, fish in 24‐h darkness deposited more energy for growth and spent less energy on respiration and excretion. Results show that photoperiod manipulation can affect growth, and that a continuous dark regime could improve growth in lenok at this stage of development.     

CAM-idling in Hoya carnosa (Asclepiadaceae)

In the leaf sueculent Asclepial Hoya carnosa (L.) R. Br., CAM photosynthesis occurred under well-watered conditions, as characterized by diurnal gas exchange and changes in titratable acidity. Following 10–12 days of severe water stress, the plants shifted from CAM to a modified CAM-idling mode of metabolism. CAM-idling was characterized by complete or almost complete stomatal closure accompanied by CAM-like diurnal changes in titratable acidity. H. carnosa plants maintained this CAM-idling mode of photosynthesis for at least 8 weeks. Upon reirrigation, the plants returned to the original CAM mode within 1 week. These results suggested that CAM-idling is a reversible, intermediate form of sustained metabolism which enables plant survival under conditions of extended drought.This work was supported in part by NSF Grant PCM 8200366 and in part by the Science and Education Administration of the United States Department of Agriculture under Competitive Grant 5901-0420-8-0018-0.     

Structure of the receptacular nectary and circadian metabolism of starch in the ant‐guarded plant Ipomoea cairica (Convolvulaceae)

Nectaries occur widely in Convolvulaceae. These structures remain little studied despite their possible importance in plant–animal interactions. In this paper, we sought to describe the structure and ultrastructure of the receptacular nectaries (RNs) of Ipomoea cairica, together with the dynamics of nectar secretion. Samples of floral buds, flowers at anthesis and immature fruits were collected, fixed and processed using routine methods for light, scanning and transmission electron microscopy. Circadian starch dynamics were determined through starch measurements on nectary sections. The secretion samples were subjected to thin layer chromatography. RNs of I. cairica were cryptic, having patches of nectar‐secreting trichomes, subglandular parenchyma cells and thick‐walled cells delimiting the nectary aperture. The glandular trichomes were peltate type and had typical ultrastructural features related to nectar secretion. The nectar is composed of sucrose, fructose and glucose. Nectar secretion was observed in young floral buds and continued as the flower developed, lasting until the fruit matured. The starch content of the subglandular tissue showed circadian variation, increasing during the day and decreasing at night. The plastids were distinct in different portions of the nectary. The continuous day–night secretory pattern of the RNs of I. cairica is associated with pre‐nectar source circadian changes in which the starch acts as a buffer, ensuring uninterrupted nectar secretion. This circadian variation may be present in other extrafloral nectaries and be responsible for full daytime secretion. We conclude that sampling time is relevant in ultrastructural studies of dynamic extranuptial nectaries that undergo various changes throughout the day.     

Diurnal Fluctuations in the Activity of the Enzyme Nitrate Reductase in Lemna paucicostata

Diurnal fluctuations were observed in nitrate reducta.se activity in Lemna paucicostatu Hegelm, grown under 16-h photoperiod. The enzyme activity showed a single peak in the middle of the photo period, i.e. 8 h after the onset of light. The activity decreased during the latter half of the photoperiod and reached a minimum level in the middle of the dark period. The peak in enzyme activity persists in continuous light as well as continuous dark, at least for a couple of days, suggesting the existence of endogenous rhythmicity     

The Morphology and Permeability of Isolated Cuticular Membranes of Hoya carnosa R. Br. (Asclepiadaceae)

This investigation is in part an extension of previous leafcuticle observations made on 52 other taxa among 34 families.Dewaxed, chemically isolated, adaxial and abaxial cuticularmembranes and transverse leaf sections of the wax-flower plant(Hoya carnosa R. Br.) were examined using ordinary stainingtechniques and light-microscopy methods. Evidence is presentedfor the existence of ubiquitous, discrete, naturally occurringcuticular pores, concomitant with anticlinally oriented trans-cuticularcanals, distributed randomly throughout the cuticular matrix.The surface of the adaxial cuticular membrane contains approx.6540 unclustered pores per mm², the abaxial approx. 4680 poresper mm². Pore and canal diameters range between 0.5 and 0.75µm. The canals are often arcuate and their lengths aredirectly related to cuticle thickness. No correlations werefound between cuticle thickness and either pore numbers or poreand canal diameters. Based upon experiments with various pHindicators, solutions, and stains, the dewaxed, dry cuticularmembrane of H. carnosa appears to be both distinctly hydrophilicand selectively permeable through a myriad of microscopicallyvisible pores and canals permeating its matrix. A de novo interpretationof gross cuticle morphology based solely upon light microscopyobservations is presented by semi-diagrammatic illustrations. Hoya carnosa R. Br., wax-flower (wax-plant), cuticular membranes, cuticular pores, transcuticular canals, permeability     

The pineal clock affects behavioral circadian rhythms but not photoperiodic induction in the Indian weaver bird (Ploceus philippinus)

We investigated whether pineal is part of the circadian clock system which regulates circadian rhythms of activity and photosensitivity in the Indian weaver bird (Ploceus philippinus). Two experiments were performed. The first experiment examined the induction of testicular growth, and androgen-dependent beak pigmentation and luteinizing hormone (LH)-specific plumage coloration in pinealectomised (pinx) and sham-operated (sham) birds exposed to short day (8 h light: 16 h darkness, 8L:16D) and long day (16L:8D) for 9 months in the late breeding and early regressive phase (October), or the late regressive and preparatory phase (January) of the annual testicular cycle. As expected, short days were non-stimulatory, and long days stimulated testicular growth, beak pigmentation and plumage coloration. There was no difference in the response between pinx and sham birds subjected to short or long days in October, but the response was enhanced in pinx birds that were subjected to long day in January. In the second experiment circadian behavioral rhythms were studied (activity pattern in singly housed birds) in weaver birds first exposed at two different phases of the annual testicular cycle to a stimulatory photoperiod (12L:12D in preparatory phase or 13L:11D in early breeding phase) and then released into dim continuous light (LL_dim). All birds showed synchronization to the light period before and after the pinealectomy; there was no difference in the response between pinx and sham birds. When released into LL_dim, sham birds exhibited circadian rhythmicity continuously, whereas pinx birds lost circadian rhythmicity after some cycles. Considered together, these results suggest that circadian clock residing within the pineal gland regulates the circadian rhythm in activity, but not the circadian rhythm involved in photoperiodic induction of the Indian weaver bird.     

Hoya aphylla (Apocynaceae,Asclepiadoideae), a new leafless species from Laos

The leafless species Hoya aphylla Aver. N.S. Khang & Averyanova (Apocynaceae) is described and illustrated as a new species for science from Hin Nam No National Protected Area of Laos (central Laos, Khammoune province). In its floral morphology, this new species may be similar to H. lyi, H. carnosa and H. multiflora, but it differs in corona color and shape of corona segments. Like the first two mentioned species, the new species may be referred to the type section of the genus, but in terms of it its leafless habit it exhibits an isolated position.     

Effects of light and circadian clock on growth and chlorophyll accumulation of Nannochloropsis gaditana

Circadian clocks synchronize various physiological, metabolic and developmental processes of organisms with specific phases of recurring changes in their environment (e.g. day and night or seasons). Here, we investigated whether the circadian clock plays a role in regulation of growth and chlorophyll (Chl) accumulation in Nannochloropsis gaditana, an oleaginous marine microalga which is considered as a potential feedstock for biofuels and for which a draft genome sequence has been published. Optical density (OD) of N. gaditana culture was monitored at 680 and 735 nm under 12:12 h or 18:6 h light‐dark (LD) cycles and after switching to continuous illumination in photobioreactors. In parallel, Chl fluorescence was measured to assess the quantum yield of photosystem II. Furthermore, to test if red‐ or blue‐light photoreceptors are involved in clock entrainment in N. gaditana, some of the experiments were conducted by using only red or blue light. Growth and Chl accumulation were confined to light periods in the LD cycles, increasing more strongly in the first half than in the second half of the light periods. After switching to continuous light, rhythmic oscillations continued (especially for OD₆₈₀) at least in the first 24 h, with a 50% decrease in the capacity to grow and accumulate Chl during the first subjective night. Pronounced free‐running oscillations were induced by blue light, but not by red light. In contrast, the photosystem II quantum yield was determined by light conditions. The results indicate interactions between circadian and light regulation of growth and Chl accumulation in N. gaditana.     

Description of the individual growth of Daphnia magna (Crustacea: Cladocera) through the von Bertalanffy growth equation. Effect of photoperiod and temperature

Daphnia magna is a cladoceran used as a model organism in aquatic ecology and ecotoxicology studies. Because growth is a critical parameter to study the effect of environmental conditions on the development of zooplankters, the somatic growth of D. magna was measured here and described by the von Bertalanffy growth equation (VBGE), a mathematical model widely used in fisheries management. For this purpose, the effect of two temperatures (20 and 25°C) and two photoperiod conditions (12:12 and 16:8, light:dark) was assayed. Experiments began with neonate females and were finished when parthenogenetic females reached the age of 41 days; they were fed the microalga Ankistrodesmus falcatus (400,000 cell ml⁻¹, 12 mg l⁻¹, dry weight). According to the VBGE, maximal length (L _max) was inversely correlated with the growth rate (K). The highest L _max (6.45 mm) was for the females grown at 20°C with the 12:12 photoperiod, whereas the maximum growth rate (K = 0.182 ± 0.010) was for individuals grown at 25°C with the 12:12 photoperiod. The number of clutches during the studied period was significantly higher for females grown at 25°C, 12:12. Temperature affected the growth rate and the maximum size in D. magna; interaction of temperature with photoperiod was also noteworthy. The VBGE was a nifty way to assess the effects of the tested environmental factors.     

The effect of photoperiod on male sexual behaviour in a north adriatic strain of the migratory locust

The Panzano strain of the migratory locust, Locusta migratoria, originating from North Italy, was used for laboratory experiments under conditions of either 12 or 16 hrs illumination per day. Preliminary experiments demonstrated that in this strain 16 hrs day-length led to delayed and reduced egg-laying, while 12 hrs day-length induced early and intensive oviposition. In subsequent experiments sexual behaviour of males kept under either regime of the photoperiod was observed in the presence of receptive females. Kept under 12 hrs day-length, males exhibited vigorous sexual behaviour, approximately at the same level of intensity from the 2nd till the 5th–6th week of the experiment. In contrast, under 16 hrs illumination the intensity of male sexual behaviour gradually increased from the 2nd till the 9th–10th week. Thus, photoperiod affected the intensity of male sexual behaviour in this strain.

---

Résumé La souche Panzano de Locusta migratoria provenant du nord de l'Italie, a été utilisée pour des expériences de laboratoire sous des conditions d'éclairement de 12 h ou 16 h par jour. Des expériences préliminaires avaient montré que chez cette souche, une durée d'éclairement de 16 h par jour retardait et réduisait la ponte, tandis qu'avec un éclairement de 12 h celle-ci était précoce et intense. Dans les expériences ultérieures on a observé le comportement sexuel de mâles, en présence de femelles réceptrices, alors qu'ils sont soumis à l'une ou l'autre de ces deux photopériodes. Maintenus sous une photopériode de 12 h les mâles montraient un comportement sexuel intense, se maintenant à peu près au même niveau de la seconde semaine jusqu'à la 5ème ou 6ème semaine de l'expérience. Au contraire, avec un éclairement de 16 h, l'intensité du comportement sexuel s'accroissait graduellement de la seconde semaine jusqu'à la 9ème ou 10ème semaine. Ainsi, la photopériode modifiait l'intensité du comportement sexuel mâle chez cette souche de Locusta migratoria.

     

Circadian rhythm of sex pheromone-releasing behaviour in females of the dermestid beetle,Trogoderma glabrum: Regulation by photoperiod

The effects on sex pheromone-releasing, or calling behaviour, of diel photoperiods of varying daylength, of light cycle phase shifts, and of continuous illumination were investigated in Trogoderma glabrum females. On light régimes with 8 to 20 hr daylengths, calling maxima tended to centre close to photophase midpoints. Although influencing the time of day at which calling occurred, daylength had little effect on the amount of activity or the length of the calling period. When 16 : 8 LD light cycles were advanced or delayed by 4 hr, the time of day at which calling peaks were observed shifted within 2 to 4 cycles so that a constant phase relationship with photoperiod was maintained. Daily calling peaks were evident in groups of females exposed to between 1 and 5 days of continuous illumination, but mean calling time occurred earlier in the day as light exposures were lengthened. It was concluded that a circadian rhythm of calling behaviour exists in T. glabrum females. and that the rhythm can be entrained to 24 hr periodicity by photoperiod.     

野牛草克隆分株酶促活性氧清除系统对差异光周期的响应

以盆栽野牛草克隆分株为材料,将克隆分株分别标记为O(姊株)和Y(妹株),设置连接组和断开组两种处理,其中,连接组中O分株和Y分株通过节间子相连,断开组则剪断分株节间子;两组处理的O分株光周期均设置为光照12h/黑暗12h,Y分株光周期均设置为黑暗12h/12h光照(恰好与O分株相反),经过7d的差异光周期处理后进行72h全光照稳定培养,并于全光照条件下在48h内连续测定各分株叶片超氧化物歧化酶(SOD),过氧化物酶(POD),过氧化氢酶(CAT),抗坏血酸过氧化物酶(APX)的活性以及丙二醛(MDA)的含量,探讨野牛草叶片酶促活性氧清除系统对差异光周期的响应特征。结果表明,差异光周期处理1周后,全光照条件下,断开状态的野牛草克隆分株O和Y间叶片中SOD、POD、CAT、APX活性以及MDA含量在24h内基本呈现相反的变化趋势,而野牛草相连克隆分株O和Y间叶片中以上指标在24h内呈现趋于一致的变化规律。研究发现,野牛草酶促活性氧清除系统活性在一天内呈现节律性表达模式,且差异光周期处理下的野牛草相连克隆分株的活性氧清除系统的活性的节律性变化趋于同步。     

Experimental evidence of masculinization by continuous illumination in a temperature sex determination teleost (Atherinopsidae) model: is oxidative stress involved?

The present study evaluates the influence of continuous light on phenotypic sex ratios in Chirostoma estor, a temperature sex determination animal model. Relative gene expression levels of 5 day old larvae were performed on two early gonad differentiation genes (sox9 and foxl2), two stress axis activation genes (gcr1 and crf) and four reactive oxygen species (ROS) antagonist effector genes (sod2, ucp2, gsr and cat). Two light treatments were applied from fertilization; control (12L:12D) simulated natural photoperiod and a continuous illumination photoperiod. By the end of the trial (12 weeks after hatching), differentiated and normal gonads were clearly identifiable in both treatments by histological observations. Regarding sex ratio, 73% of phenotypic males were found in continuous illumination compared with 40% in controls. Consistently, the sox9 gene (involved in early testis differentiation) showed an over expression in 64% of the individual larvae analysed compared with foxl2 (ovarian differentiation) suggesting a masculinization tendency in continuous illumination. On the other hand, only 36% of individuals showed the same tendency in the control treatment consistent with phenotypic sex ratios found under normal culture conditions. Relative gene expression results did not show significant difference in sod2, ucp2 and gcr1 levels, but cat, gsr and crf showed significantly higher expression levels in the continuous illumination treatment suggesting that both, the stress axis and ROS response mechanisms were activated at this time. This study suggests, a link between continuous light, oxidative stress and environmental sex determination in vertebrates. However, further research is necessary to describe this possible upstream mechanism that may drive some aspects of sexual plasticity in vertebrates.