Endophyte transmission and activity in the Anabaena-Azolla association

Summary The survival of Azolla was studied in an artificial system which simulated the soil/water interface and the desiccation of soil during a fallow period in lowland rice culture. Tests with non-sporulating and sporulating Azolla fronds showed that Azolla only survives with sporulated fronds. At their reappearance the Azolla fronds already harboured the Anabaena endophyte. A detailed light microscopic and transmission electron microscopic study of macro- and micros-porocarp formation and development revealed that the endophyte is transmitted by the macrosporocarps and not by the microsporocarps. The Anabaena cells within the macrosporocarps are found just below the indusium cap. These cells are not nitrogen-fixing akinetes. The free-living Anabaena cells at the stem apex and below the overarching developing leaves do not bear heterocysts and accordingly are non nitrogen-fixing. During the development of the leaf the Anabaena enters the leaf cavity, but later the pore of this, cavity closes and the imprisoned cyanobacteria are lysed before the leaf decays. As the Azolla leaves age a nitrogen-fixing capability is successively built up concomittantly with the production of heterocysts. Heterocyst frequencies of 40–50% can be found inAnabaena azollae. Usually a gradient of nitrogen-fixing capacity occurs along the Azolla rhizome with two distinct peaks at leaf number 7/8 and at leaf number 13/14 from the apex.     

Metabolism and translocation of fixed nitrogen in the nodulated legume

Summary Nitrogen (N₂) fixed by Rhizobium bacteroids in the legume nodule is excreted as ammonia to the surrounding host cell where it is efficiently assimilated into the amide group of glutamine. Generally glutamine is a minor exported solute of nitrogen, being further metabolised to asparagine in temperate species and to the ureides, allantoin and allantoic acid in tropical species. These solutes serve as the principal translocated forms of nitrogen in xylem. Compartmentalisation of the pathways of nitrogen metabolism and the role of ammonia in regulation of their activity is examined in nodules of both asparagine-forming (Lupinus albus L.) and ureide-forming (Vigna unguiculata L. Walp) symbioses.     

Selection for components related to body composition in mice: direct responses

Summary Replicated within full-sib family single-trait selection was conducted for 10 generations in mice for (1) high or low 12-week epididymal fat pad percentage (100 x epididymal fat pad weight/body weight) or (2) high or low 12-week hind carcass percentage (100 x hind carcass weight/body weight). Pooled realized heritabilities based on high, low and divergent selection were 0.66±0.09, 0.65±0.13 and 0.66±0.05 for epididymal fat pad percentage and 0.48±0.08, 0.33±0.08 and 0.40±0.04 for hind carcass percentage. The pooled realized genetic correlation (r_{G R}) between epididymal fat pad percentage and hind carcass percentage based on divergence was –0.67±0.04. Other estimates of (r_{G R}) were: epididymal fat pad percentage with body weight (0.57±0.05); epididymal fat pad percentage with epididymal fat pad weight (1.17±0.05); hind carcass percentage with body weight (–0.61±0.09); hind carcass percentage with hind carcass weight (–0.05±0.11). Indirect measures of fat and lean tissue percentages were highly heritable, and (r_{G R}) between them would be desirable from the standpoint of analogous types of traits in livestock. In the same context, undesirable (r_{G R})'s were found between epididymal fat pad percentage and body weight and between hind carcass percentage and body weight.Paper No. 10957 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, North Carolina 27695-7601, USA. The use of trade names in this publication does not imply endorsement by the North Carolina Agricultural Research Service of the products named, nor criticism of similar ones not mentioned     

Cultivar and pH effects on competition for nodule sites between isolates of Rhizobium in beans

Two experiments were carried out to evaluate the effect of acidity on bean-Rhizobium competition for nodule sites. SevenPhaseolus vulgaris host cultivars differing in acid-pH tolerance were grown in sand culture, and irrigated using a sub-irrigation system and nutrient solutions of pH 4.5, 5.0, 5.5, and 6.0. A mixed inoculant of two antibiotically markedRhizobium leguminosarum bvphaseoli strains CIAT899 (acid-tolerant) and CIAT632 (acid-sensitive) was used. The acid-tolerant CIAT899 dominated CIAT632 in nodule occupancy across all cultivars and pH treatments. Although several of the varieties had previously been identified as PH-tolerant, and these cultivars performed better than those reported to be acid sensitive, all showed a marked increase in nodulation and plant development when the pH was raised from 4.5 to 6.0. The second experiment using a modified Leonard jar system varied the inoculation ratio between CIAT899 and UMR1116 (acid-sensitive, inefficient in N₂-fixation) and contrasted nodulation response for the bean varieties Preto 143 (pH-tolerant) and Negro Argel (pH-sensitive) at 3 pH treatments (4.5, 5.5, 6.5). There was a significant effect of host cultivar, ratio of inoculation, and pH on the percentage of nodule occupancy by each strain. At low pH CIAT899 had higher nodule occupancy than UM1116 in the variety Negro Argel but had the same percentage of nodulation when the variety was Preto 143. Increasing the cell concentration of UMR1116 produced more inefficient nodules at all treatment combinations and reduced plant growth for both cultivars used.     

TheRhizobium-legume symbiosis Two methods to discriminate between nodules and other root-derived structures

Summary Two methods have been developed in order to discriminate between lateral roots, nodules and root-derived structures which exhibit both root and nodule histological features and which can develop on legumes inoculated with certainRhizobium mutants. The first method, known as the clearing method, allows the observation by light microscopy of cleared undissected root-structures. The second, known as the slicing method, is a complementary technique which provides a greater degree of structural information concerning such structures. The two methods have proved invaluable in defining unequivocally the nature of the interaction between a rhizobial strain and a legume host.     

Voltage clamping with single microelectrodes: Comparison of the discontinuous mode and continuous mode using the Axoclamp 2A amplifier

Summary The voltage clamp technique is a powerful method for studying the physiology of excitable membrane. This technique has made possible the determination of ionic responses generated by activation of either receptor-mediated or voltage-dependent processes. The development of the whole-cell, tight-seal voltage clamp method has allowed the analysis and examination of membrane physiology at the single cell level. The method allows the characterization of voltage-dependent ionic conductances both at the macroscopic (whole-cell) and at the microscopic (unitary conductance or single channel) level in cells less than 10 µm in diameter, a feat difficult to achieve with conventional fine-tipped micropipettes.In this paper, several methologies used for culturing neuronal and non-neuronal cells in the laboratory are described. A comparison between the two modes of voltage clamp using blunt-tipped patch-microelectrodes, the switching (discontinuous) and the non-switching (continuous) modes, of the Axoclamp-2A amplifier is made. Some results on membrane currents obtained from neuronal and non-neuronal cells using the single electrode whole-cell tight-seal voltage clamp is illustrated. The possible existence of two inactivating K⁺ currents, one dependent on Ca⁺⁺ the other is not, is discussed.     

Adventitions shoot formation on excised leaves of in vitro grown shoots of apple cultivars

Leaves taken from micropropagated shoots of several apple (Malus domestica Borkh.) cultivars were cultured in vitro on Linsmaier & Skoog (LS) medium or the rice anther culture medium of Chu et al. (N6) containing various concentrations of either benzyladenine (BA) or thidiazuron (TDZ) plus naphthaleneacetic acid (NAA). Of the TDZ concentrations tested, 10 M was most effective and it was equivalent to, or better than, 22 M BA for both the percentage of leaves regenerating shoots and number of shoots formed per regenerating leaf in almost every experiment. Lower concentrations of NAA (1.1 and 5.4 M) gave best results with both BA and TDZ. N6 medium gave consistently better results than LS. Lowering total salt concentration or total N concentration of LS to that of N6 did not improve the response nor did changing the NO₃:NH₄ ratio. The 3–4 leaves on the most distal part of the shoot were most responsive and tended to form the most adventitious shoots. Placing the leaf cultures in the dark for the first 2–3 weeks of the culture period produced the best results. Optimum results were obtained by culturing leaves from the distal part of the shoot in the dark for 2 weeks on N6 medium containing 10 M TDZ and 1.1 or 5.4 M NAA, then moving the cultures to 16 h daylight at a photon flux of 60 mol s^-1m^-2.     

In vitro propagation of green-foliaged Dracaena fragrans Ker.

Callus tissue was induced in young stem segments cultured on MS based media supplemented with 0.25–0.5 mg l^-1 2, 4-D. Shoots were differentiated on media containing 0.5–1.0 mg l^-1 BA and 0.5–2.0 mg l^-1 IBA or 0.1–0.2 mg l^-1 NAA. The same media were suitable for shoot multiplication. Shoot elongation and rooting were strongly inhibited by BA and stimulated by auxins IBA and NAA. Medium containing 0.5 mg l^-1 IBA was optimal for rooting. Root elongation was stimulated by light and inhibited in darkness. Transfer of rooted plantlets to outdoor conditions was feasible and special hardening procedures were not required. Among more than 5000 plants produced by this procedure only 9 off-type plants with variegated leaves were found.     

An immunohistochemical study on the postnatal development of rat nasal-associated lymphoid tissue (NALT)

Summary This study concerns the development of nasal-associated lymphoid tissue in the rat, using immuno- and enzyme-histochemical staining techniques on cryostat sections. Nasal-associated lymphoid tissue is present at birth as a small accumulation of mainly T lymphocytes and non-lymphoid cells; B cells are rare. Distinct areas of T and B cells appear at 10 days after birth; by that time high endothelial venules are also observed. Intra-epithelial lymphocytes are present, most of them being T-helper cells. ED1+ macrophages are seen throughout the tissue. The proportion of ED1+cells does not change during ontogeny. ED2+cells (tissue macrophages) are present predominantly at the border between the lymphoid tissue and the surrounding connective tissue, in all age-groups. ED3+mononuclear cells are scattered throughout the nasal-associated lymphoid tissue of young animals. Later on, the ED3+ cells migrate into the border-area between lymphoid and connective tissue. Ia+ non-lymphoid cells in the nasal lymphoid tissue increase in number during ontogeny. Only a few of them show acid phosphatase activity, indicating that the proportion of classical scavenger macrophages is low. Some of them may be antigen presenting (dendritic) cells. Ia+ dendritic cells also occur between the epithelial cells. Moreover, some epithelial cells express the Ia marker.     

Methanol metabolism in thermotolerant methylotrophic Bacillus strains involving a novel catabolic NAD-dependent methanol dehydrogenase as a key enzyme

The enzymology of methanol utilization in thermotolerant methylotrophic Bacillus strains was investigated. In all strains an immunologically related NAD-dependent methanol dehydrogenase was involved in the initial oxidation of methanol. In cells of Bacillus sp. C1 grown under methanol-limiting conditions this enzyme constituted a high percentage of total soluble protein. The methanol dehydrogenase from this organism was purified to homogeneity and characterized. In cell-free extracts the enzyme displayed biphasic kinetics towards methanol, with apparent K _m values of 3.8 and 166 mM. Carbon assimilation was by way of the fructose-1,6-bisphosphate aldolase cleavage and transketolase/transaldolase rearrangement variant of the RuMP cycle of formaldehyde fixation. The key enzymes of the RuMP cycle, hexulose-6-phosphate synthase (HPS) and hexulose-6-phosphate isomerase (HPI), were present at very high levels of activity. Failure of whole cells to oxidize formate, and the absence of formaldehyde-and formate dehydrogenases indicated the operation of a non-linear oxidation sequence for formaldehyde via HPS. A comparison of the levels of methanol dehydrogenase and HPS in cells of Bacillus sp. C1 grown on methanol and glucose suggested that the synthesis of these enzymes is not under coordinate control.Abbreviations RuMP ribulose monophosphate - HPS hexulose-6-phosphate synthase - HPI hexulose-6-phosphate isomerase - MDH methanol dehydrogenase - ADH acohol dehydrogenase - PQQ pyrroloquinoline, quinone - DTT dithiothreitol - NBT nitrobluetetrazolium - PMS phenazine methosulphate - DCPIP dichlorophenol indophenol