Parturition and alate morph determination in the aphid Megoura viciae

In reproducing apterae of Megoura viciae, parturition is often completely arrested during periods of isolation from the host plant. In contrast, surgical removal of the rostrum (including the stylets), amputation of the extremities of the legs, or decapitation, all stimulate parturition away from the plant. These operations also induce alata-producing aphids to revert immediately to the production of apterae, but have no detectable effect on aptera-producers. Carbon dioxide or ether anaesthesia and nitrogen narcosis have a similar action on this maternally controlled response. Although the rostrum and tibio-tarsus bear sensilla whose removal might well be involved in inducing parturition, the influence on morph change is probably indirect and is to a great extent associated with the delay in the resumption of parturition. The effect can be reproduced by isolating individual aphids away from the food plant. The morph change cannot, however, be attributed to starvation since it also occurs when the genital pore of an actively feeding aphid is temporarily occluded. The change in physiology appears to be associated with the retention of embryos at a time when there is no sensory input from crowding.

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Résumé Chez les Megoura viciae aptères, la parturition est souvent complètement arrêtée pendant les périodes de séparation de la plante hôte. Par opposition, l'amputation chirurgicale du rostre (y compris les stylets), de l'extrémité des pattes (tarse et une partie du tibia), ou la décapitation, stimulent toutes la parturition en l'absence de la plante. Les pucerons induits à produire des ailés (élevage antérieur en groupe) retournent, après ces opérations, immédiatement vers la production d'aptères. Le dioxyde de carbone ou l'anesthésie à l'éther et la narcose à l'azote ont une action semblable sur ce déterminisme maternel. Bien que l'élimination des sensilla portées par le rostre et par l'ensemble tarse-tibia puisse être déterminante dans l'induction de la parturition, l'action sur le changement de type semble être principalement associée au retard consécutif dans la reprise de la parturition. Les anesthésiants qui, eux aussi, retardent l'apparition ou la reprise de la parturition, ont probablement une action indirecte du même type.Les pucerons groupés, isolés de la plante hôte pendant plus de 24 h, ont aussi tendance à retourner immédiatement à la production d'aptères. Ce changement de type ne peut, cependant, être attribué au jeûne puisqu'il se produit aussi quand, chez un puceron s'alimentant activement, le pore génital est momentanément bouché. Le changement physiologique semble être associé à la rétention des embryons à un moment où il n'y a pas l'influence sensorielle du groupement.Aucun de ces traitements, à l'exception du groupement, n'induit des pucerons antérieurement isolés à devenir des producteurs d'ailés.

     

The putative brain photoperiodic photoreceptors in the vetch aphid,Megoura viciae

In an attempt to identify the brain photoreceptors that mediate the photoperiodic response of the vetch aphid, Megoura viciae, we utilised immunocytochemical techniques and employed 20 antibodies directed against invertebrate and vertebrate opsins and phototransduction proteins. A sub-set of these antibodies (to Drosophila rhodopsin 1: RH1-1; vertebrate cone opsins: COS-1; CERN-874; CERN-933; vertebrate rod opsin: CERN-901; vertebrate arrestin: AB-Arr; vertebrate transducin+arrestin+rhodopsin kinase+cGMP phosphodiesterase: CERN-911; and vertebrate cellular retinoid binding protein: CRALBP) consistently labelled an anterior ventral neuropile region of the protocerebrum. These anatomical findings, coupled with previous localised illumination and micro-lesion studies, provide strong evidence that this region of the aphid brain houses the photoperiodic photoreceptors. The present study also confirms that the medial (Group I) neurosecretory cells are not the photoperiodic photoreceptors.     

The effects of starvation on development and reproductive potential of apterous virginoparae of vetch aphid, Megoura viciae

Apterous virginoparae of Megoura viciae Buckton starved during the fourth instar lost weight but nevertheless completed their development, showing that growth and development could proceed independently. Their weight at the end of starvation was a function of initial weight. Aphids used considerable amounts of lipid during starvation, and if allowed to feed on becoming adult showed a delayed onset to reproduction. Starvation in the fourth instar arrested embryogenesis, and in small aphids resulted in degeneration of gonadal and somatic tissues.

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Résumé Des virginipares de M. viciae Buckton, privées de nourriture à partir de la mue au quatrième stade, achèvent leur développement et muent en adulte en perdant du poids; elles montrent que croissance et développement peuvent être indépendants. Le jeûne pendant le quatrième stade provoque en moyenne la perte de 84% des réserves lipidiques. Le poids à la mue imaginale dépend significativement du poids lors de la mue du quatrième stade: de petits pucerons achèvent leur développement plus lentement que les gros. Le jeûne pendant le 4ème stade interrompt l'ovulation et le développement des gonades; lorsque les adultes peuvent se réalimenter après ce jeûne, la période précédant la ponte est plus longue. L'examen histologique montre que les pucerons les plus petits subissent une régression somatique et gonadale plus importante que les plus lourds: la dégenérescence des gonades apparaît chez les petits pucerons après le jeûne au 4ème stade, tandis que les plus gros voient leur potentiel reproductif réduit par un arrêt de l'embryogenèse.

     

Responses of male aphids to the female sex pheromone in Megoura viciae Buckton

Oviparae of Megoura viciae Buckton release a sex pheromone from numerous plaques on their hind tibiae. Adult males react to this scent in a variety of ways: inactive males are aroused; males already walking orient to the odour source; and copulatory activity is increased. These responses are only evoked within a few centimetres of the stimulus source and decline rapidly as the stimulus is diluted. Male responsiveness to the pheromone is unchanging throughout the light phase (of LD 12 : 12), although they show a marked circadian locomotor rhythm. Their sensitivity develops early in adult life and thereafter remains high, decreasing only slightly before death. However, although very young adults will orient to a pheromone source, they do not show copulatory responses until they are sexually mature. Larval males neither respond to the pheromone nor possess the antennal receptors (secondary rhinaria) which are essential for the adult reaction. Mating, and attempted copulation with suitable objects (e.g. coloured beads), is dependent upon appropriate visual and tactile stimuli; it can occur in the absence of the pheromone but is enhanced by its presence. Habituation to the scent does not prevent copulation, and conversely, copulation does not affect subsequent responsiveness to the pheromone. Once copulation has begun the pheromone has no effect on the efficiency of insemination. Vigorous intergeneric responses to the pheromones of female M.viciae and Acyrthosiphon pisum Harris were observed.     

Untersuchungen zur Stoffwechselphysiologie von Megoura viciae Buckt., einer phloemsaugenden Aphide

Ohne ZusammenfassungTeil einer Dissertation der Naturwissenschaftlichen Fakultät der Universität Würzburg. Meinem Lehrer, Herrn Priv.-Doz. Dr. W. Kloft, danke ich für die Übertragung des Themas, Herrn Prof. Dr. K. Gösswald für die Überlassung eines Arbeitsplatzes. Die Arbeit wurde durch Mittel der Deutschen Forschungsgemeinschaft gefördert, welche Herrn Priv.-Doz. Dr. Kloft zur Verfügung standen.     

Ultrastructural features of egg development in oviparae of the vetch aphid, Megoura viciae buckton

In the ovaries of the oviparous morph of the aphid, Megoura viciae, resting oocytes are located in the basal region of each germarium. During previtellogenic egg development, electron-dense spheres appear in the ooplasm. During vitellogenesis a brush border develops at the oolemma, and numerous protein and lipid-like spheres accumulate in the egg cytoplasm. Follicle cells are of two morphologically distinct types, termed 'type 1' and 'type 2' follicle cells. Unlike the more numerous 'type 1' cell, 'type 2' cells do not become patent. The acellular tunica propria exterior to follicle cell apices remains intact throughout egg development. During late vitellogenesis symbiont invasion of eggs takes place via 'receptor' cells encircling the pedicel at the posterior egg pole. These cells shrink and/or degenerate to create intercellular spaces that facilitate symbiont transmission. The end of vitellogenesis is marked by vitelline membrane formation and secretion of the chorionic layers, at which time the next egg in the ovariole undergoes final stages of previtellogenic growth and enters vitellogenesis.     

Chromosomal abnormalities in males-producing eggs: a study in Megoura viciae (Homoptera,Aphididae)

Pairing stages of homologous chromosomes suggesting the occurrence of an endomeiotic process during the maturation of parthenogenetic oocytes have been found in Megoura viciae. This study underlines the importance of these synaptic stages as far as the determination of the male sex is concerned. Maturation divisions with 8 chromosomes (6 univalents + 2 bivalents) instead of 9 chromosomes (1 bivalent XX + 8 autosomes) have been found in males-producing eggs. The presence of these two bivalents causes aneuploidy in male embryos and probably abortion.     

The influence of natural enemies on wing induction in Aphis fabae and Megoura viciae (Hemiptera: Aphididae)

Previous studies have shown that the aphid species, Aphis fabae Scopoli and Megoura viciae Buckton, do not produce winged offspring in the presence of natural enemies, in contrast to results for the pea aphid (Acyrthosiphon pisum (Harris)) and the cotton aphid (Aphis gossypii Glover); but these studies did not involve exposing aphids directly to natural enemies. We exposed colonies of both A. fabae and M. viciae to foraging lacewing (Chrysoperla carnea (Stephens)) larvae and found that the predators did not induce winged morphs among offspring compared to unexposed controls. Colonies of A. fabae responded to an increase in aphid density with increasing winged morph production, while such response was not found for M. viciae. We suggest that different aphid species differ in their susceptibility to natural enemy attack, as well as in their sensitivity to contact.     

Cytological and electrophoretic analysis of DNA methylation in the holocentric chromosomes of Megoura viciae (Homoptera, Aphididae).

Chromosomal and purified DNA methylation patterns were determined in the holocentric chromosomes of Megoura viciae by treatment with MspI and HpaII. Both enzymes produced a clear C-like banding pattern but widely digested one telomere of the X chromosome, which appeared as heterochromatic after C-banding treatment and brightly fluorescent after chromomycin A3 staining. Quantitative microfluorometric evaluations of DNA extraction performed on cytological preparations showed that both isoschizomers resulted in the same DNA extraction (about 30%). Contrary to what was found by in situ endonuclease treatment, the electrophoretic patterns of purified and digested DNA showed that digestion with MspI was slightly more extensive than that with HpaII in a zone of fragments ranging from 23 to 9 kb. This result indicates that aphid chromatin is not wholly unmethylated. The discrepancy between electrophoretic and cytological data has been explained by taking into consideration that DNA fragments with high molecular weights could be cleaved in situ by the enzymes but not extracted from the chromatin.     

Occurrence of nitric oxide synthase in Megoura viciae Buckton (Homoptera, Aphididae): an histochemical and immunohistochemical localisation

Nitric oxide (NO) is known to be involved in many physiological reactions of insects. We analysed NOS localisation in aphids of the species Megoura viciae by means of histochemical reaction for the NADPH-diaphorase activity and immunohistochemical methods for uNOS, nNOS and iNOS. The obtained data provided a complex and peculiar pattern of NOS distribution in cells and tissue of M. viciae. The histochemical reaction for NADPH-diaphorase was an indicative, but not exact marker of NOS localisation in aphids. The use of anti uNOS antiserum (frequently applied in insects) was of limited value in our specimens, whereas more satisfactory results were obtained with anti nNOS and iNOS antisera of human origin. The results of Western blot analysis confirmed the immunohistochemical ones, showing an aphid protein that reacted strongly with the polyclonal antibody anti-iNOS and anti-nNOS while a similar protein band was weakly immunoreactive with the polyclonal antibody anti-uNOS. Our results suggest that NO, prevalently synthesised by calcium/calmodulin-dependent isoform, plays important physiological roles both in adult and embryological stages of aphids. The data of principal interest was NOS presence in bacteriocytes, cells that host symbiotic prokaryotes belonging to the species Buchnera aphidicola, and in nuclei of adipocytes and gut cells.     

Biochemical effects of molecular crowding

Cell cytoplasm contains high concentrations of high-molecular-weight components that occupy a substantial part of the volume of the medium (crowding conditions). The effect of crowding on biochemical processes proceeding in the cell (conformational transitions of biomacromolecules, assembling of macromolecular structures, protein folding, protein aggregation, etc.) is discussed in this review. The excluded volume concept, which allows the effects of crowding on biochemical reactions to be quantitatively described, is considered. Experimental data demonstrating the biochemical effects of crowding imitated by both low-molecular-weight and high-molecular-weight crowding agents are summarized.Translated from Biokhimiya, Vol. 69, No. 11, 2004, pp. 1522–1536.Original Russian Text Copyright © 2004 by Chebotareva, Kurganov, Livanova.     

Macromolecular crowding and size effects on probe microviscosity

Development of biologically relevant crowding solutions necessitates improved understanding of how the relative size and density of mobile obstacles affect probe diffusion. Both the crowding density and relative size of each co-solute in a mixture will contribute to the measured microviscosity as assessed by altered translational mobility. Using multiphoton fluorescent correlation spectroscopy, this study addresses how excluded volume of dextran polymers from 10 to 500 kDa affect microviscosity quantified by measurements of calmodulin labeled with green fluorescent protein as the diffusing probe. Autocorrelation functions were fit using both a multiple-component model with maximum entropy method (MEMFCS) and an anomalous model. Anomalous diffusion was not detected, but fits of the data with the multiple-component model revealed separable modes of diffusion. When the dominant mode of diffusion from the MEMFCS analysis was used, we observed that increased excluded volume slows probe mobility as a simple exponential with crowder concentration. This behavior can be modeled with a single parameter, β, which depends on the dextran size composition. Two additional modes of diffusion were observed using MEMFCS and were interpreted as unique microviscosities. The fast mode corresponded to unhindered free diffusion as in buffer, whereas the slower agreed well with the bulk viscosity. At 10% crowder concentration, one finds a microviscosity approximately three times that of water, which mimics that reported for intracellular viscosity.     

Long-term effects of crop management on Rhizobium leguminosarum biovar viciae populations

Little is known about factors that affect the indigenous populations of rhizobia in soils. We compared the abundance, diversity and genetic structure of Rhizobium leguminosarum biovar viciae populations in soils under different crop managements, i.e., wheat and maize monocultures, crop rotation, and permanent grassland. Rhizobial populations were sampled from nodules of pea- or vetch plants grown in soils collected at three geographically distant sites in France, each site comprising a plot under long-term maize monoculture. Molecular characterization of isolates was performed by PCR-restriction fragment length polymorphism of 16S-23S rDNA intergenic spacer as a neutral marker of the genomic background, and PCR-restriction fragment length 0polymorphism of a nodulation gene region, nodD, as a marker of the symbiotic function. The diversity, estimated by richness in types and Simpson's index, was consistently and remarkably lower in soils under maize monoculture than under the other soil managements at the three sites, except for the permanent grassland. The highest level of diversity was found under wheat monoculture. Nucleotide sequences of the main rDNA intergenic spacer types were determined and sequence analysis showed that the prevalent genotypes in the three maize fields were closely related. These results suggest that long-term maize monoculturing decreased the diversity of R. leguminosarum biovar viciae populations and favored a specific subgroup of genotypes, but the size of these populations was generally preserved. We also observed a shift in the distribution of the symbiotic genotypes within the populations under maize monoculture, but the diversity of the symbiotic genotypes was less affected than that of IGS types. The possible effect of such changes on biological nitrogen fixation remains unknown and this requires further investigation.     

Some remarks on molecular biology

The binding energy of a very long molecular chain, composed of different classes of molecules, depends in general on the order of the molecules. It is shown that under very general conditions there exists for a givenbrutto chemical composition of a chain, a class of chains which is characterized by a total binding energy which is equal to the total binding energy of any other prescribed chain of different composition within the limits of unsharpness of the energy level. This establishes a criterion formapping of a class of configurations of long chain molecules on another class. To the extent that a mapping constitutes a generalized code those results contribute to the theory of molecular codes. Applying to our results the results of a previous paper (1959,Bull. Math. Biophysics,21, 309–326), we arrive at the conclusion that the self-replication of a living molecule may be the property not of a particular structure but of classes of structures.     

Molecular crowding effects on structure and stability of DNA

Living cells contain a variety of biomolecules including nucleic acids, proteins, polysaccharides, and metabolites as well as other soluble and insoluble components. These biomolecules occupy a significant fraction (20-40%) of the cellular volume. The total concentration of biomolecules reaches 400gL(-1), leading to a crowded intracellular environment referred to as molecular crowding. Therefore, an understanding of the effects of molecular crowding conditions on biomolecules is important to broad research fields such as biochemical, medical, and pharmaceutical sciences. In this review, we describe molecular conditions in the cytoplasm and nucleus, which are totally different from in vitro conditions, and then show the biochemical and biophysical consequences of molecular crowding. Finally, we discuss the effect of molecular crowding on the structure, stability, and function of nucleic acids and the significance of molecular crowding in biotechnology and nanotechnology.     

Measuring the effects of macromolecular crowding on antibody function with biolayer interferometry

ABSTRACT

Biotherapeutic proteins are commonly dosed at high concentrations into the blood, which is an inherently complex, crowded solution with substantial protein content. The effects of macromolecular crowding may lead to an appreciable level of non-specific hetero-association in this physiological environment. Therefore, developing a method to characterize the diverse consequences of non-specific interactions between proteins under such non-ideal, crowded conditions, which deviate substantially from those commonly employed for in vitro characterization, is vital to achieving a more complete picture of antibody function in a biological context. In this study, we investigated non-specific interactions between human serum albumin (HSA) and two monoclonal antibodies (mAbs) by static light scattering and determined these interactions are both ionic strength-dependent and mAb-dependent. Using biolayer interferometry (BLI), we assessed the effect of HSA on antigen binding by mAbs, demonstrating that these non-specific interactions have a functional impact on mAb:antigen interactions, particularly at low ionic strength. While this effect is mitigated at physiological ionic strength, our in vitro data support the notion that HSA in the blood may lead to non-specific interactions with mAbs in vivo, with a potential impact on their interactions with antigen. Furthermore, the BLI method offers a high-throughput advantage compared to orthogonal techniques such as analytical ultracentrifugation and is amenable to a greater variety of solution conditions compared to nuclear magnetic resonance spectroscopy. Our study demonstrates that BLI is a viable technology for examining the impact of non-specific interactions on specific biologically relevant interactions, providing a direct method to assess binding events in crowded conditions.     

The biology of Peronospora viciae on pea: the development of local and systemic infections and their effect on vining yield

The development of infection by Peronospora viciae (Berk.) Casp. was studied in field and plot experiments during 1967, 1968 and 1969. Numbers of primary systemically infected plants appeared shortly after emergence and reached a maximum by the end of May. Such plants acted as primary infectors and did not survive more than 3 weeks. Maximal sporangial release occurred between 06.00 and 08.00 h BST with a peak at 07.00 h, after 1–2 h insolation. Inoculation of leaves at 10.30 h resulted in greater infection than inoculation at 20.00 h. The result suggested a possible maturation requirement of sporangia following release from the sporangiophore. Initially the spread of foliar (local lesion) infection in the crop was sporadic and localized in cvs. Jade, Gregory, Dark Skinned Perfection (D.S.P.) and Frühe Kleine Pfälzerin (F.K.P.) during the 3 years. The number of plants of all cvs. with foliar infection reached 100% by mid-June. Lesions rarely exceeded 15% of the leaf and stipular area of the mature plant. No effect of foliar lesions, at the highest level of infection, in haulm or seed fresh weight could be detected in cvs. F.K.P., Jade and D.S.P. Secondary systemic infection occurred sporadically in field crops usually with dense haulm growth. Plants of Jade with systemic infection were rated according to the number of nodes affected. There was a small but significant increase in haulm and seed fresh weight in plants with up to two nodes affected. But plants with three to five nodes showed a progressive reduction in the yield of green seed (resulting from a reduction in the number of podding nodes) and in pod size and haulm fresh weight. Total losses in Jade during 1968 and 1969 with all grades of infection did not exceed 4 % of the green seed crop. Systemic infection in some plants resulted in the death of the apex and the production of healthy, normally podding tillers. Such plants had no commercial value due to delayed maturation. Pod infection in Jade occurred largely independently of systemic symptoms: the overall level during 1969 did not exceed 16 % and only 2 % were severely infected. Total losses (from both systemic and pod infection) in Jade during 1968 and 1969 represented only a negligible part of the crop.