Rough and fine linkage mapping of the Rhizobium meliloti chromosome.

Summary A circular linkage map of the Rhizobium meliloti chromosome, obtained from R68.45-mediated crosses, has been revised by cotransductional analysis, after general transduction by DF2 phage.Three short chromosomal regions have been mapped by cotransduction. Comparison between conjugal and cotransductional data suggests that R68.45-mediated linkage measures are indeed rough. Cotransduction seems to be a useful tool for improving the linkage map of R. meliloti.     

Infection of sugar cane by the nitrogen-fixing bacterium Acetobacter diazotrophicus

Significant nitrogen fixation has recently been demonstratedin Brazilian sugar cane (Saccharum officinarum) cultivars knownto form associations with a number of diazotrophs, includingAcetobacter diazotrophicus, an acid-tolerant endophytic bacteriumwhich grows best on a sucrose-rich medium. In a series of experiments,aseptically-grown sugar cane plantlets were rooted in a liquidmedium and inoculated with A. diazotrophicus originally isolatedfrom field-grown sugar cane. After 4, 7, 9, and 15 d, plantswere examined under light, scanning and transmission electronmicroscopes and the presence of A. diazotrophicus on and withinplant tissues was confirmed by immunogold labelling. By 15 d,external bacterial colonization was seen on roots and lowerstems, particularly at cavities in lateral root junctions. Theloose cells of the root cap at root tips were a site of entryof the bacteria into root tissues. Both at lateral root junctionsand root tips, bacteria were also seen in enlarged, apparentlyintact, epidermal cells. After 15 d, bacteria were present inxylem vessels at the base of the stem, many connected via mucusto spiral secondary thickening. There was no obvious pathogenicreaction to the bacteria within the xylem. From these observations,it is proposed that, under experimental conditions, A. diazotrophicusfirstly colonized the root and lower stem epidermal surfacesand then used root tips and lateral root junctions to enterthe sugar cane plant where it was distributed around the plantin the transpiration stream. It is further suggested that thexylem vessels in the dense shoots of mature plants are alsoa possible site of N₂-fixation by diazotrophs as they providethe low pO₂ and energy as sucrose necessary for nitrogenaseactivity. Key words: Acetobacter diazotrophicus, endophyte, infection, nitrogen fixation, sugar cane.     

Herbaspirillum, an endophytic diazotroph colonizing vascular tissue 3Sorghum bicolor L. Moench

Leaves of Sorghum bicolor were examined at 5 d and 14 d afterinoculation with the N₂-fixing endophytic bacteria Herbaspirillumseropedicae and Herbaspirillum rubrisubalbicans. Plants inoculatedwith H. rubrisubalbicans expressed symptoms of ‘red stripedisease’ i.e. red stripes along the secondary veins ofthe leaf blade close to the inoculation point and spreadingup the leaves. Infected leaves showed dense colonization byH. rubrlsubalbicans in regions showing red stripe symptoms at5 d after inoculation. The infection was confined within thevascular system, in particular, the protoxylem and associatedlacunae, which were often completely filled with bacteria, withsome of the latter expressing nitrogenase. The bacteria wererecognized using H. rubrisubalbicans-speciflc antibodies andimmunogold labelling, which also showed that the antibody reactedwith material on the surface of the bacteria, and that thismucus was released into the lumen of the xylem. At 14 d afterinoculation, disease symptoms were slightly more severe, withboth meta- and protoxylem being even more heavily colonizedin parts of the leaf showing red stripes. However, a stronghost defence response was also apparent at this stage, withgums lining the walls of the vessels and enclosing the bacteria,although the latter were still actively dividing. At the edgesof visible disease symptoms, plant gums filled the xylem; bacteriahad formed distinct colonies within these gums, with some ofthe colonies associated with the xylem walls. Plants inoculatedwith H. seropedicae either did not express the disease or showedvery mild symptoms close to the inoculation point. In the lattercase, H. seropedicae were localized within protoxylem vesselsand the metaxylem was partly occluded with plant-derived gums.By contrast with H. rubrisubalbicans, H. seropedicae was alsolocalized in leaves at 14 d without disease symptoms and didnot always appear to elicit a host response, i.e. they colonizedthe walls of metaxylem, with the xylem vessels themselves remainingunoccluded and largely free of gums. The fine line separatingplant pathogens, endophytes and symbioses is discussed in lightof these results. Key words: Herbaspirillum, Sorghum bicolor, nitrogen fixation, endophyte, xylem     

Intrinsic labeling of bovine milk with enriched stable isotopes of zinc

Bovine milk was labeled intrinsically with enriched stable isotopic zinc for human bioavailability studies. Intrajugular administration of zinc isotopes temporarily increased the plasma zinc concentration of Ayrshire cows by as much as 76%, but milk zinc concentration and the distribution of zinc between casein and whey did not change appreciably. Milk zinc isotopic enrichment reached 105 and 613 atom % excess for 67Zn and 70Zn, respectively within 4-12 hr of zinc administration and decreased gradually over several days. This degree of isotopic enrichment is sufficient for testing bioavailability to infants of intrinsic zinc from milk-based formulas.     

A simple screening procedure for adenylate kinase, hexokinase and glucose-6-phosphate dehydrogenase deficiencies

A simple screening procedure for the detection of adenilate kinase (AK), hexokinase (Hx) or glucose-6-phosphate dehydrogenase (G6PD) deficiencies in blood, is described. It consists of two assays : in the first, the ATP formed by blood AK is coupled to Hx and G6PD, and in the second, the glucose-6-phosphate formed by blood Hx is coupled to G6PD. The enzyme activities are visually estimated by the reduction of NADP+ (non-fluorescent) to NADH (fluorescent). The appearance of fluorescence in the first assay indicates that the three enzyme activities are present. The absence of fluorescence could be due to the deficiency of any one of the three enzymes; in this case the second assay used in combination with the Beutler's screening test for G6PD permits the detection of the specific enzymatic deficiency.     

Water and nutrient relations of woody perennials from tropical dry forests

Abstract. Dry forests occupy a larger area in the tropics than rain forests. They grow under a wide range of rainfall conditions; the determining characteristics are the occurrence of a dry season of 2–6 months duration, and the dominance of deciduous woody perennials. The knowledge of the ecophysiological properties of woody perennials from these forests, essential for the development of forest restoration strategies, is still scanty. This paper describes the ecophysiological behaviour of 10 species of woody perennials growing in a secondary dry forest, which has been recovering since 1944 on the hills of the Botanical Garden of Caracas. Four species, Astronium graveolens, Bauhinia megalandra, Sapindus saponaria and Tabebuia chrysantha, were initially planted while the others, Bursera simaruba, Capparis flexuosa, Erythroxylon cumanense, E. densum, E. orinocense and Eugenia casearioides, reproduced from seed sources existing in forest remnants growing nearby. Specific leaf areas measured are relatively high, covering a range from 11 to 34 m²/kg. Nitrogen and phosphorus levels are also high compared to data reported from dry forests elsewhere, therefore no nutrient limitation for photosynthetic productivity is apparent. Osmotic pressure (π) of leaf sap, extracted from frozen samples taken during the growing season, is correlated with the corresponding content of soluble sugars and the ions Mg and K; Ca-ions do not play a significant role in explaining the variance of π. Most species maintained a diluted leaf sap during the rainy season, characterized by π values between 5–15 bar. The osmotic pressure increased strongly in older leaves and during the dry season. Four species showed more stable π values throughout the growing season, with C. flexuosa and B. simaruba characterized by higher and lower π values, respectively. Proline was found to be a reliable indicator of water stress in these woody species, the amount of proline measured in leaf sap being logarithmically correlated with the corresponding π value. Osmotic pressure of leaf sap and leaf xylem tension was higher during the dry season for all species, while the contrary was true for leaf conductance. Leaf conductance was better correlated with leaf-air vapour pressure deficit than with leaf xylem tension. The most drought-tolerant species were C. flexuosa, E. casearioides and the three Erythroxylon species. Drought resistance of B. simaruba, B. megalandra and A. graveolens was associated with their high sensitivity to leaf-air VPD and lower leaf conductances. The other two species occupied intermediate positions.     

Stability and transmissibility of the cryptic plasmids of Rhizobium meliloti GR4

Rhizobium meliloti strain GR4 harbours two cryptic plasmids sharing extensive regions of homology between them and with other non-symbiotic plasmids of different strains of R. meliloti. They both are very stable showing a segregation rate of less than 0.1% loss per generation. pRmeGR4a (115 MDa) is a self-transmissible plasmid at a variable frequency to other species, and it is also responsible for promoting, at low frequency, the contransfer of pRmeGR4b (140 MDa), the other cryptic plasmid of the strain. A 4.8 kb PstI fragment of pRmeGR4a, responsible for the high stability in cis of this plasmid, has been isolated and several recombinant plasmids have been constructed showing different segregation rates in the strains used in this study. Their stabilities can be considerably improved by insertion of the stabilization mrs/par region of RK2.     

Location of nif genes on large plasmids in Rhizobium strains isolated from legume tree root nodules

Abstract The presence of plasmids in Rhizobium strains isolated from legume tree root nodules has been studied. Bacteria isolated from Acacia melanoxylon, A. cyanophylla, Prosopis chilensis and Sophora microphylla harbour 2 to 5 plasmids of an M _r higher than 115 · 10⁶. Hybridization experiments have shown DNA homology between plasmid pID1 ( R. meliloti 41 nifH and D genes) and one of the plasmids in each of the bacteria studied.