Structure and Organization of Effective Peanut and Cowpea Root Nodules Induced by Rhizobial Strain 32H1

A comparison of the structure and organization of nodular tissuesand bacteroids of peanut and cowpea induced by Rhizobium sp.strain 32H1 was madc 4 to 5 weeks after inoculation when nitrogenaseactivity reaches the peak. Observations revealed major differencesthat may have a role in the different rates of nitrogen fixationshown by the two species. All cell types in cowpca nodules werelarger than those of peanut. The inner cortex of cowpea hadan ‘endodermis-like’ layer of cells which was absentin peanut. All cells in the bactcroidai zone of peanut wereinfected but in cowpca many remained free of bactcroids. Thebacteroid containing cells of peanut were isodianietrical anduniform in size with a central vacuole and a nucleus surroundedby tightly arranged bactcroids enclosed singly in peribacteroidalmembrane sacs. Such cells in cowpea were mostly elongated witha nucleus and one or more vacuoles. The bacteroids within cowpeacells were arranged without any particular order with more spacefor host cellular material. They were mostly present singlyin peribacteroidal membrane sacs which sometimes fused to enclosemore than one bactcroid. The hosts seem to play the dominantrole in the differentiation of nodular tissue and the morphogenesisof bacteroids in symbiotic systems induced by the same strainof Rhizobium. Key words: Peanut, Cowpea, nodule structre     

Phytohormones, Rhizobium Mutants, and Nodulation in Legumes : III. Auxin Metabolism in Effective and Ineffective Pea Root Nodules

High specific activity [³H]indole-3-acetic acid (IAA) was applied to the apical bud of intact pea (Pisum sativum L. cv Greenfeast) plants. Radioactivity was detected in all tissues after 24 hours. More radioactivity accumulated in the nodules than in the parent root on a fresh weight basis and more in effective (nitrogen-fixing) nodules than in ineffective nodules (which do not fix nitrogen).

For most samples, thin layer chromatography revealed major peaks of radioactivity at the R_F values of IAA and indole-3-acetylaspartic acid (IAAsp) and further evidence of the identity of these compounds was obtained by chromatography in other systems. Disintegrations per minute due to IAA per unit fresh weight were significantly greater for root than for nodule tissue, but were not significantly different for effective and ineffective nodules. Radioactivity due to IAAsp, expressed both on a percentage basis and per unit fresh weight, was significantly greater for nodule than for root tissue and significantly greater for the effective nodules than for the ineffective nodules. When [³H]IAA was applied to effective nodules, IAAsp was the dominant metabolite in the nodule. The data suggest that metabolism of auxins may be important for the persistence of a functional root nodule.

     

Formation of Nitrosylleghemoglobin in Nodules of Nitrate-Treated Cowpea and Pea Plants

The formation of nitrosylleghemoglobin (LbNO) was examined incowpea and pea nodules in relation to the inhibition of nitrogenfixation by nitrate. Leghemoglobin was of the ferrous type andwas mainly converted to LbNO in cowpea nodules when the acetylene-reducingactivity decreased to 45% of control values as a result of thesupply of nitrate. In nodules of nitrate-treated pea plants,leghemoglobin was also of the ferrous type and LbNO was a minorcomponent of leghemoglobin. The levels of LbNO isolated fromnodules corresponded to the levels of LbNO calculated from equilibriumconstants for LbNO and the concentration of nitrite in nodules.The dissociation rate constants for LbNO from both cowpea andpea were much smaller than those for LbO₂ or LbCO, as is alsothe case in soybean. These results indicate that the inhibition of the functionsof leghemoglobin, due to the accumulation of LbNO, induces adecrease in nitrogen fixation in cowpea nodules, and that theinhibition of nitrogen fixation in pea nodules is not relatedto the formation of LbNO. (Received July 2, 1990; Accepted October 9, 1990)     

Metabolism under Microaerobic Conditions of Mitochondria from Cowpea Nodules

A method is described for isolating mitochondria from nodules of cowpea (Vigna unguiculata [L.] Walp.) under completely anaerobic conditions. The mitochondria were immediately active when incubated aerobically with substrates, and their respiration rates were higher than mitochondria prepared in air. The mitochondria lacked fumarate reductase and were not inhibited by 5% CO₂. When incubated under microaerobic conditions, their respiration could be measured by leghemoglobin spectroscopy. Microaerobic respiration was inhibited approximately 50% by 1 millimolar malonate, and was completely inhibited by cyanide. O₂ uptake and the ATP/O ratio declined under microaerobic conditions, and therefore ATP production may be low in the environment of infected nodule cells.     

富钾基因型烤烟根系形态和生理特性研究

在水培条件下,以富钾基因型烤烟'农大202'和一般型品种'NC89'、'净叶黄'为材料,研究了不同供钾水平下烟草根系形态和生理特性的基因型差异.结果显示:与低钾水平相比,各品种烤烟在常钾水平下的根系鲜重、体积、干重、总吸收面积、活跃吸收面积、比表面积、根系活力、H+分泌量、阳离子交换量(CEC)、可溶性蛋白和ATPase活性均明显升高,但根系比表面积有所下降,并以'农大202'烤烟的变化幅度较大;在常钾水平下,'农大202'表现出良好的根系形态和较强的生理特性,但其在低钾胁迫下却低于一般型品种,耐低钾能力较弱.研究表明,烟草的根系形态和生理特性及其对环境供钾水平的响应存在明显的基因型差异;提高环境供钾水平能明显促进各基因型烟草根系的形态发育和生理活性,从而增强其对钾的吸收、转运能力.     

Heat Shock Tolerant Mutants of Rhizobium meliloti Forming Ineffective Nodules

Six heat shock tolerant mutants of Rhizobium meliloti Rmd201 were isolated through transposon Tn5 mutagenesis. The symbiotic assays of these mutants with alfalfa plants, showed four of these mutants to be affected in nitrogenase effectivity also. These four mutants could be classified into two separate complementation groups hssA and hssC through R-prime mediated merodiploid constructions. The hssC mutant Rmd1040 also showed poor interaction with phages indicating surface alterations. The results indicated possible involvement of these loci in symbiosis as well as heat shock response.     

烟草磷效率的基因型差异及其与根系形态构型的关系

以17个具有代表性的主要烟草基因型为材料,通过盆栽试验和培养基栽培试验,研究烟草磷效率的基因型差异及其与根系形态构型的关系,为磷高效烟草品种选育提供理论依据.结果表明,施磷肥能够显著增加各供试烟草基因型的生物量及氮、磷和钾的累积量;供试烟草的磷效率和氮、钾累积量存在显著基因型差异,土壤盆栽试验中,低磷条件下的'云烟85'生物量和磷累积量分别是'NC82'的4.06倍和3.34倍,氮和钾累积量分别是'K358'的4.06倍和3.75倍;供试烟草可划分为磷低效低产型、磷低效高产型、磷高效高产型、磷高效低产型等4种类型,其中的'云烟85'、'K326'、'云烟2号'、'RG11'和'红花大金元'属于磷高效高产型,是现代磷高效高产品种选育的理想材料.供试烟草基因型的根系形态构型与其磷效率显著相关,与磷低效低产型烟草'G28'和'许金1号'相比,磷高效高产型烟草'云烟85'和'K326'在高低磷条件下根系均较发达,总根长和根表面积均较大;磷有效性对烟草根构型具有调节作用,在缺磷条件下,磷高效基因型具有浅根根构型,而磷低效基因型具有深根根构型.     

Cowpea Rhizobia Producing Dark Nodules: Use in Competition Studies

During a program of screening rhizobia from West Africa, it was found that some strains produced nodules of unusually dark appearance on cowpeas, but not on peanuts, soybeans, pigeon peas, or mung beans. The dark pigmentation was in the bacteroid zone, was not correlated with nodule effectiveness, and was additional to the leghemoglobin pigment. Only rhizobial strains with a nongummy (“dry”) colony morphology produced dark nodules. Visually distinguishable pink and dark nodules formed on the same root when a mixture of pink and dark strains was applied as inoculum. The dark-nodule phenotype was therefore appraised as a marker and found to be useful for studying nodulation competition with strains of the orthodox pink-nodule type. The competitiveness of 10 pink-nodule strains was examined relative to a black-nodule strain, IRc 256; a range of competitiveness was obtained of less competitive than, equally competitive to, or more competitive than IRc 256. Patterns of primary (early) nodulation were generally the same as patterns of secondary (later) nodulation. Mixed infections by dark and pink strains produced piebald nodules, the frequency of occurrence of which was much greater among primary than among secondary nodules.     

3种荒漠盐生植物根系及根毛形态特征的比较研究

在水培条件下,针对梭梭、囊果碱蓬和钠猪毛菜3种荒漠盐生植物,研究它们苗期在不同盐浓度条件下根系和根毛形态的差异。结果表明：一定浓度的盐分可以促进3种盐生植物生长,但较高浓度的盐抑制其生长,特别是对根系生长的抑制作用更大。在同样盐浓度下,钠猪毛菜的生长最快,生物量也最大。在盐分浓度较低时,3种盐生植物的主根长和总根长都有所增加,与对照相比,囊果碱蓬增加的幅度较大,但高浓度的盐会抑制根系总长度的增加,其中囊果碱蓬较梭梭和钠猪毛菜抑制的程度轻。盐分对3种植物的根系平均直径没有显著的影响,但有减小的趋势。在水培条件下,梭梭和囊果碱蓬的根系上、中、下部分布的较均匀,而钠猪毛菜的根系中部比上部和下部有显著的增加,盐分对每种植物的根系的分布没有显著的影响。3种植物的根毛与根系生长环境中的盐浓度没有明显相关性;3种植物之间,根毛的长度和密度有显著差异。从根系和根毛的形态特征可以推断：囊果碱蓬比梭梭和钠猪毛菜具有较强的抗旱性和耐极薄能力;梭梭的耐盐能力较其它2种植物差,囊果碱蓬的耐盐性最强。     

The Effect of Ageing on the Leghemoglobin of Cowpea Nodules

Experiments were conducted to study the effect of ageing of the host and senescence of the nodules on the content and composition of the proteins of leghemoglobin in nodules of cowpea (Vigna sinensis L.). Total and heme protein contents of leghemoglobin were the highest in the red nodules and dwindled with the onset of nodule senescence. In the fully senescent green nodules the total proteins decreased to a very low level, while the heme proteins were almost absent. Disc electrophoretograms of leghemoglobin obtained from red and brown nodules showed nine bands out of which only four gave positive test for heme. Greening of the nodules resulted in a considerable reduction in the intensity of the bands. Senescence of the host plant resulted in a reduction in the number of leghemo-globin proteins in all types of nodules and brought about a drastic change in their electrophoretic mobility.     

Cytokinins in Root Nodules of Phaseolus mungo

Four cytokinins have been separated from extracts of root nodulesof Phaseolus mungo by thin-layer chromatography. Their activitywas determined on the basis of their ability to induce betacyaninsynthesis in cotyledons of Amaranthus caudatus. Zeatin and itsriboside showed greater activity than N⁶ (²-(isopentenyl)) aminopurine and its riboside in the bioassay. Phaseolus mungo, mung bean, cytokinins, isopentenyl, amino-purine, zeatin, betacyanin synthesis, Amaranthus caudatus     

Proteolysis during Development and Senescence of Effective and Plant Gene-Controlled Ineffective Alfalfa Nodules

Plant-controlled ineffective root nodules, conditioned by the in1 gene in Medicago sativa L. cv Saranac, undergo premature senescence and have reduced levels of many late nodulins. To ascertain which factors contribute to premature senescence, we have evaluated proteolysis as it occurs throughout the development of ineffective Saranac (in1Sa) and effective Saranac nodules. Cysteine protease activities with acidic pH optimum and enzyme proteins were present in both genotypes. We found that acidic protease activity was low in effective Saranac nodules throughout their development. In contrast, by 2 weeks after inoculation, acid protease activity of in1Sa nodules was severalfold higher than that of Saranac nodules and remained high until the experiment was terminated 8 weeks later. This increase in protease enzyme activity correlated with an increase in protease protein amounts. Increased protease activity and amount in in1Sa nodules was correlated with a decrease in nodule soluble protein. The time at which in1Sa nodules initially showed increased protease activity corresponded to when symbiosis deteriorated. High levels of phosphoenolpyruvate carboxylase (PEPC) protein were expressed in effective nodules by 12 d after inoculation and expression was associated with low proteolytic enzyme activity. In contrast, although PEPC was expressed in in1Sa nodules, PEPC protein was not found 12 d after inoculation and thereafter. Acidic protease from in1Sa nodules could also degrade purified leghemoglobin. These data indicate that premature senescence and low levels of late nodulins in in1Sa nodules can be correlated in part with increased proteolysis.     

Reexamination of the Intracellular Localization of de Novo Purine Synthesis in Cowpea Nodules

Sucrose and Percoll density gradient centrifugation were used to separate organelles from the central zone tissue of cowpea (Vigna unguiculata L. Walp. cv Vita 3: Bradyrhizobium strain CB 756) nodules. Enzyme activity analysis has shown that both plastids and mitochondria have a full complement of enzymes for de novo purine synthesis. In vitro activities of individual component enzymes (glycinamide ribonucleotide synthetase, EC 6.3.4.13; glycinamide ribonucleotide transformylase, EC 2.1.2.2; aminoimidazole ribonucleotide synthetase, EC 6.3.3.1; aminoimidazole carboxamide ribonucleotide transformylase, EC 6.3.2.6; and adenylosuccinate-AMP lyase, EC 4.3.2.2) as well as of the whole purine pathway (from ribose-5-phosphate to inosine monophosphate) were similar in the two organelles. No significant cytosolic or bacteroidal activity of any of the purine pathway enzymes was detected on assay. These findings are contrary to earlier studies (M.J. Boland, K.R. Schubert [1983] Arch Biochem Biophys 220: 179-187; B.J. Shelp C.A. Atkins, P.J. Storer, D.T. Canvin [1983] Arch Biochem Biophys 224: 429-441) that concluded that enhanced expression of purine synthesis in nodules of ureide-forming species is localized to plastids. Significantly increased recovery of activity of key pathway enzymes (particularly of labile aminoimidazole ribonucleotide synthetase) coupled with improved assay methods and the use of Percoll in addition to sucrose for gradient centrifugation have together contributed to much higher reaction rates and more definitive analyses of particulate fractions.     

贵州福泉分布桃花水母的形态特征及分子鉴定

2017年10月,在贵州省福泉县发现了桃花水母(Craspedacusta)。观察其形态结构,福泉的桃花水母与索氏桃花水母(C. sowerbyi)高度相似。采用PCR和DNA测序技术扩增和测定了福泉采集桃花水母的核糖体小亚基rRNA基因(18SrRNA)、核糖体RNA基因内转录间隔区(ITS)及线粒体细胞色素氧化酶亚基基因(COI),并与GenBank上已有的桃花水母18S rRNA、ITS、COI基因序列进行了比对。分子鉴定结果显示,福泉的桃花水母18SrRNA、ITS、COI基因序列与索氏桃花水母相似度分别为100%、92%、99%,确定贵州省福泉市发现的桃花水母样品在种分类水平上为索氏桃花水母。并就桃花水母的某些形态学分类指标的标准、不同伞径之间的差值是否能作为桃花水母形态分类的一个新指标这些问题做了讨论。     

mRNA of Leghemoglobins from Alfalfa Root Nodules

This paper described the methods of isolation, purification, and in vitro translation of alfalfa leghemoglobin mRNA. It was pointed out that the mRNA of alfalfa leghemoglobin accounts for about 1.4% of total RNA, and 24% of poly (A) +-mRNA of alfalfa nodule. This mRNA has 6, 9, 26S value populations with different molecular weights of about 0.15×106, 0.28×106, 0.48×l06 respectively. These S value mRNAs were coded for different components of leghemoglobin.     

Characteristics of Modified Leghemoglobins Isolated from Soybean (Glycine max Merr.) Root Nodules

Hemoprotein derivatives of an abundant soybean (Glycine max Merr.) root nodule leghemoglobin, Lba, were studied for their modified spectral characteristics and physical properties. Three modified hemoprotein derivatives of Lba (Lbam1, Lbam2, and Lbam3) were purified by preparative isoelectric focusing. The ferric forms of these pigments were green and exhibited anomalous spectra in the visible region as compared to the Lba3+ forms. These modified pigments showed a hypochromic shift of 10 nm for the charge transfer absorption maximum; however, differences were not apparent in the Soret region. Upon binding with nicotinate, the [alpha] and [beta] bands were shifted significantly into the red region as compared to the Lba3+ nicotinate complex. The three Lbam fractions were reduced by dithionite or by NADH in the presence of riboflavin. Lbam2+ also bound nicotinate and displayed absorption spectra indistinguishable from those of Lba2+ nicotinate. In contrast to Lba2+, Lbam2+ displayed aberrant spectra when bound with either O2 or CO. These complexes exhibited a prominent charge transfer band at approximately 620 nm and failed to exhibit spectra characteristic of Lba2+O2 and Lba2+CO. The protein moiety of these modified pigments was intact because their tyrosine/tryptophan ratios and their amino acid compositions were identical with those of Lba, nor were differences observed in the peptide profiles resulting from trypsin digests of purified Lba and Lbams. Automated Edman degradation of selected peaks further confirmed the intactness of the protein backbone including the absence of deamination. Pyridine hemochromogen for heme from Lbams could be formed, and the spectra displayed distinct differences compared to those of Lba. A new peak at 580 nm and a loss of a peak at 480 nm were observed for all three Lbams.     

Pathways of Nitrogen Assimilation in Cowpea Nodules Studied using N(2) and Allopurinol

In the presence of 0.5 millimolar allopurinol (4-hydroxypyrazolo [3,4-d]pyrimidine), an inhibitor of NAD:xanthine oxidoreductase (EC 1.2.3.2), intact attached nodules of cowpea (Vigna unguiculata L. Walp. cv Vita 3) formed [¹⁵N]xanthine from ¹⁵N₂ at rates equivalent to those of ureide synthesis, confirming the direct assimilation of fixed nitrogen into purines. Xanthine accumulated in nodules and was exported in increasing amounts in xylem of allopurinol-treated plants. Other intermediates of purine oxidation, de novo purine synthesis, and ammonia assimilation did not increase and, over the time course of experiments (4 hours), allopurinol had no effect on nitrogenase (EC 1.7.99.2) activity. Negligible ¹⁵N-labeling of asparagine from ¹⁵N₂ was observed, suggesting that the significant pool (up to 14 micromoles per gram of nodule fresh weight) of this amide in cowpea nodules was not formed directly from fixation but may have accumulated as a consequence of phloem delivery.