Quantitative cytology of the alfalfa generative cell and its relation to male plastid inheritance patterns in three genotypes

Summary Studies utilizing restriction analysis of plastid DNA, as well as those employing chlorophyll-deficient mutants, have shown a high frequency of paternal plastid transmission in alfalfa. Recent research has also shown that plastid inheritance patterns among alfalfa genotypes and are under genetic control. In a previous study we were unable to detect any correlations between qualitative, three-dimensional ultrastructure of generative cells and male plastid transmission strength in certain genotypes. In the present study we used serial ultrathin sectioning, computerized reconstruction and quantitation, and stereology to further analyze generative cells within mature pollen. Measurements included volumes and surface areas of cells, nuclei, and organelles, as well as organelle number and distribution. Three genotypes were investigated, one that is a strong transmitter of male plastids (genotype 301), one that is a weaker transmitter of male plastids (genotype 7W), and a third that is an even weaker male plastid transmitter (genotype MS-5). Our results show that genotype MS-5 has significantly fewer plastids/generative cell than either of the other genotypes, which may account for it being a relatively poor transmitter of male plastids. However, plastid number does not explain known differences in male plastid inheritance between genotypes 301 and 7W, since plastid number does not differ significantly between these two genotypes. Regarding the other features of generative cells measured in this study, no consistent correlations were found that might account for differences in male plastid inheritance patterns between genotypes. Plastid distribution is equal in each end of the spindle-shaped generative cell in all genotypes studied. Similar relative results were found with regard to mitochondria within generative cells; however, comparative genetic data are not available on mitochondrial transmission patterns in alfalfa genotypes.     

Lotus japonicus, an autogamous, diploid legume species for classical and molecular genetics

In the Leguminosae plant family, few of the individual plant species have been used for plant molecular biology research. Among the species investigated no obvious representative ‘model’ legume has emerged. Here a member of the tribe Loteae, Lotus japonicus (Regel) Larsen is proposed as a candidate. L. japonicus is a diploid, autogamous species, with a good seed set, and a generation time of approximately 3 months. The haploid genome consists of six chromosomes and the genome size was estimated to be relatively small (0.5 pg per haploid complement). L. japonicus is susceptible to Agrobacterium tumefaciens and transgenic plants can be regenerated after hygromycin or kanamycin selection. Tissue culture conditions and procedures for transformation and regeneration are described. Stable transformation is demonstrated by segregation of the hygromycin selectable marker after selfing of transgenic plants or test crosses. The possibility of mapping polymorphic DNA markers inbred lines of L. japonicus is also discussed.     

Resistance in the wild crop relatives Avena macrostachya and Hordeum bogdani to the aphid Rhopalosiphum padi

In previous screening tests the two wild crop relatives Avena macrostachya (Bal., ex Coss. et Dur.) and Hordeum bogdani (Wil.) demonstrated a high degree of resistance to the aphid Rhopalosiphum padi (L.). In a choice situation using wild and cultivated oats and barley, alate aphids settled in lower numbers on the wild species. The results were, however, variable in the Avena combination. Nymph production was significantly higher, development time shorter and adult weight higher on the cultivated varieties. From the third instar and onwards the excretion of honeydew was significantly lower on the resistant plants. In general the honeydew contained less than 1% free amino acids although excreta from H. vulgare contained 3.5%. The percentage of free amino acids found in the honeydew was similar for all plant species (5.2–7.6%) except for H. vulgare, on which the aphids excreted 22% of the amounts ingested. Amino acids excreted in high proportions on all plants included asparagine, -aminobutyric acid, glutamic acid, and glycine. Tissue sectioning did not reveal any obvious mechanical barriers to stylet penetration. The potential use of these wild species as sources for aphid resistance breeding in oats and barley is considered.

---

Résumè Lors d'examens systématiques antérieurs, Avena macrostachya (Bal. ex Coss. & Dur.) et Hordeum bogdani (Wil.) ont présenté une résistance élevée au puceron Rhopalosiphum padi (L.). Lorsqu'ils avaient un choix comprenant de l'avoine et de l'orge cultivés, les pucerons ailés ont atterri en nombres moins importants sur les espèces sauvages. Les résultats étaient cependant variables dans le complexe avoine. La production de nymphes et le poids des adultes étaient plus élevés sur espèces cultivées, ainsi que la durée du développement était plus longue sur les espèces sauvages. A partir du troisième stade, l'excrétion de miellat a été significativement plus faible sur les espèces résistantes. En général, le miellat y contenait moins de 1% d'acides aminés bien que sur H. vulgare il en contînt 3,5%. Les pourcentages d'acides aminés libres du miellat étaient semblables sur toutes les plantes (5,2–7,6%), à l'exception de H. vulgare sur lequel les pucerons excrétaient 22% des taux ingérés. Les acides aminés excrétés en fortes quantités sur les différentes plantes, comprenaient l'asparagine, l'acide -aminobutyrique, l'acide glutamique et la glycine. Des coupes de tissus n'ont révélé aucun obstacle mécanique clair à la pénétration des stylets. Les possibilités d'utiliser ces espèces sauvages comme source de résistance aux pucerons dans la sélection de l'avoine et de l'orge ont été examinées.

     

Cowpea aphid performance and behaviour on two resistant cowpea lines

In a series of laboratory and climate chamber tests we compared the growth and behaviour of Aphis craccivora on one susceptible (ICV-1) and two aphid-resistant (ICV-11 and ICV-12) cowpea lines. The aphids' growth rates were much lower on the resistant cowpea lines than on the susceptible one, indicating strong antibiosis. In addition, the aphids invariably settled in higher numbers on the susceptible line than on either of the resistant. Compared to ICV-1, damaged leaves of the resistant line ICV-12 were settled upon to a higher degree than undamaged leaves, and leaf discs from the same line were even less resistant.On resistant lines individual aphids waited a significantly longer time before making their first test probe. Total probing time as well as the time preceding a decision to stay or leave was also longer.These results are discussed in relation to the possible mechanisms involved, and we also consider the effects of previous leaf feeding on the expression of resistance in the field.

---

Résumé Au cours d'expériences au laboratoire et en chambres climatisées nous avons comparé la croissance et le comportement de A. craccivora sur une lignée sensible (ICV-1) et deux lignées résistantes (ICV-11 et ICV-12) de V. unguiculata. Les vitesses de croissance des pucerons ont été beaucoup plus lentes sur les lignées résistantes que sur la lignée sensible, ce qui révèle une forte antibiose. De plus, les pucerons atterrissent invariablement en plus grand nombre sur la variété sensible. Par comparaison avec ICV-1, les atterrissages sur lignée résistante ICV-12 étaient plus nombreux sur les feuilles endommagées que sur les feuilles intactes; les disques de feuilles de cette même lignée étaient encore moins résistants.Les pucerons ont séjourné individuellement un temps plus long sur les lignées résistantes avant de faire leur premier sondage. Le temps consacré aux sondages ainsi que le temps précédant de choix entre départ ou maintien sur la feuille étaient plus longs avec les lignées résistantes.Ces résultats ont été discutés en fonction des mécanismes impliqués. Nous avons aussi examiné les effets de la consommation antérieure sur les manifestations de la résistance dans la nature.

     

A Bioluminescence Method for the Measurement of l-Glutamate: Applications to the Study of Changes in the Release of l-Glutamate from Lateral Geniculate Nucleus and Superior Colliculus After Visual Cortex Ablation in Rats

We have developed a rapid, simple, specific, and very sensitive bioluminescence method for the measurement of L-glutamate (L-Glu). Oxidation of L-Glu by glutamate dehydrogenase has been coupled with bacterial FMN reductase and luciferase. Light production (i.e., peak height or integral) was linear from less than 0.5 to 500 pmol of L-Glu. Potential interfering substances that may be encountered in brain tissue have been identified. The most potent inhibitors were ascorbate and the biogenic amines. Procedures that conferred long-term stability of the reagent mixture (greater than 8 h) were established. Bioluminescence analysis of L-Glu content in brain tissue extracts, fractions from release experiments, and human CSF corroborated respective results obtained by HPLC analysis. In this study, we have applied the method to monitor changes in the KCl-evoked release of endogenous L-Glu from milligram amounts of brain tissue, i.e., from lateral geniculate nucleus and superior colliculus after visual cortex ablation.     

Haemagglutinating and Hydrophobic Properties of Corynebacterium (Eubacterium) Suis

Corynebacterium (Eubacterium) suis strains from boars and sows haemagglutinated erythrocytes of different animal species (calf, guinea pig, poultry, pig, and human). The haemaigglutination was man nose resistant (MR) and was neither inhibited by L-fucose nor D-galactose. The hydrophobicity measured by salt aggregation test (0.1–0.9 mol/1 (NH4)2SO4) and the hydrophobic interaction chromatography test (90 % retention in octyl sepharose) together with the haemagglutinating activity, indicated the presence of fimbriae on the bacteria. The haemagglutinating and hydrophobic properties were heat-sensitive (60°C for 10 min) suggestive of the presence of a protein structure. Two types of fimbria-tion were demonstrated by electron microscopy. Fetuin and glyco^ protein inhibited the haemagglutination, whereas porcine mucin was without any effect. These results indicate that branched glycoproteins might be important receptors for these fimbriae. The pathogenic aspects of C. suis are discussed, based on recent acquired knowledge of the effect of other pyelonephritogenic bacteria.     

Quantitative ultrastructural analysis of barley sperm

Summary Complete serial ultrathin sections of seven sperm pairs, computer-assisted measurements of cell, nuclear and organelle surface areas and volumes, and three-dimensional imagery were used to demonstrate that a process of cytoplasm and organelle elimination occurs during sperm maturation in barley. The number of mitochondria per sperm cell is reduced by 50%; sperm cell surface area and volume are reduced by 30% and 51% respectively. Mean volume and surface area per mitochondrion are significantly less in mature sperms. No examples of mitochondrial fusion or degeneration were observed within sperm cells. These data, along with observations of plasma membrane apposition and vesiculation within cytoplasmic extensions containing mitochondria, support the proposition that cytoplasm and organelle loss results primarily from the formation of cytoplasmic projections that are subsequently discarded from the sperm cell body. Comparisons of the quantitative data, including the number of mitochondria, indicate that differences between sperm cells of a pair are absent to very slight. Spatial organization within the pollen grain is such that the mature sperms, as well as the sperms and vegetative nucleus, are not in close proximity.     

Receptor dynamics of closely related ligands: "fast'' and "slow'' interferons.

Two related human alpha interferons with 83% homology in their primary sequences show a similar specific activity on nonhuman cells, but a striking difference on human cells, on which alpha-1 shows 1-5% of the specific molar activity displayed by alpha-2. Both interferons were labelled with 125I, and their binding kinetics followed on growing cultures of the human Burkitt line Daudi. Binding of alpha-1 showed slower rates of association and faster rates of dissociation implying that differences in apparent binding affinity were responsible for the differences in specific molar activity. However, binding was shown to reach steady-state rather than an equilibrium, so differences in the dynamics of the ligand-receptor complexes may represent amplification of differences in the initial binding constant. alpha-2, but not alpha-1, induces a marked loss of binding sites leading to a high affinity steady-state binding. Inhibition of cell multiplication by both interferons depends on a continued stimulation by free ligands at steady-state. It is proposed that the differences in specific molar activity are, in the main, kinetic and cause alpha-1 and alpha-2 to behave respectively as "slow' and "fast' interferons.     

Predation and potential transfer of pollen in a population of Saxifraga hirculus

Body surface pollen load and gut pollen (pollen diet load) of flower visitors in a Danish population of Saxifraga hirculus were analyzed. The four most frequent visitors, Eurimyia lineata (Syrphidae), Neoascia tenur (Syrphidae), Asindulum nigrum (Mycetophilidae), and Zygaena trifolii (Zygaenidae) had total surface loads (and S. hirculus loads) of 1031 (771), 137 (106), 108 (78), and 318 (27) pollen grains, respectively. Other syrphid visitors had on average a load of 407 (192) pollen grains. Males of E. lineata, N. tenur , and A. nigrum carried more surface pollen than the females; females of E. lineata had a lot of pollen from Ranunculus acris and S. hirculus and males had a lot from S. hirculus . Females of N. tenur carried pollen mostly from S. hirculus, Lotus uliginosus and Galium uliginosum ; males carried pollen mostly from S. hirculus . The surface pollen load of A. nigrum was dominated by S. hirculus pollen, whereas that of Z. trifolii was dominated by Cirsium palustre and Lychnis flos-cuculi pollen. The amount and kind of pollen varied enormously both intraspecifically and interspecifically. Eurimyia lineata, N. tenur and other syrphids had total pollen diet loads (and S. hirculus pollen diet loads) of approximately 14900 (657), 3739 (2327) and 17784 (13040). The average syrphid contained pollen from five plant species, only one to two, however, were quantitatively important. The average diet load in females was nine times that in males in both E. lineata and N. tenur. Eurimyia lineata ate a lot of R. acris; N. tenur ate a lot of S. hirculus and G. uliginosum . Generally, no correlation was observed between size or season and the composition of surface load and diet load. Eurimyia lineata made most of the pollen visits. Thus, the seed set from this Danish population of S. hirculus was dependent on a single syrphid species.     

Quantitative three-dimensional study on the position of the female gametophyte and its constituent cells as a prerequisite for corn (Zea mays) transformation

Summary The position of the embryo sac in the spikelet and of the embryo sac's constituent cells within the sporophytic tissues of Zea mays was localized by scanning electron microscopy, serial thick sectioning, and computer three-dimensional reconstruction. Within certain limits, the embryo sac is consistently oriented in the same position inside of the spikelet. This information is a prerequisite for successful microinjections into the in situ female cells of Zea mays.