Localization of oleosomes in the root nodules of alfalfa (Medicago sativa)

1998 ◽  
Vol 76 (8) ◽  
pp. 1418-1423 ◽  
Author(s):  
A Darryl Martin ◽  
Arya K Bal ◽  
David B McKenzie
Keyword(s):  
Medicago Sativa ◽  
Vascular Tissue ◽  
Rhizobium Meliloti ◽  
Root Nodules ◽  
Hexane Extraction ◽  
En Bloc ◽  
Cortical Cells ◽  
Storage Organs ◽  
Vascular Parenchyma ◽  
Meliloti Strain

Histological studies of rhizobia-induced nodules and spontaneous pseudonodules of alfalfa were undertaken to reveal the distribution pattern of oleosomes. Alfalfa (Medicago sativa L. cv. 'Algonquin') seedlings were inoculated with Rhizobium meliloti (strain Balsac), and grown in controlled conditions. Seedlings were also grown aseptically to obtain spontaneous pseudonodules. Nodule slices were fixed in a paraformaldehyde-glutaraldehyde mixture and processed for embedding in Spurr's medium after OsO4 treatment and en bloc staining, with p-phenylenediamine (pPD) in 70% ethanol, for oleosomes. As a control, hexane extraction of lipids was employed. In pseudonodules, oleosomes were present in the vascular parenchyma, the meristem region and in the five to seven layers of cortical cells adjacent to the vascular tissue. The allocation of energy stored as triacylglycerides, in the form of oleosomes, and starch indicates that pseudonodules are a sink for carbon, possibly functioning as storage organs. In mature nodules, oleosomes were distinguished clearly in nodule parenchyma but were absent in infected cells. Young nodules were devoid of oleosomes. The prevalence of oleosomes in the mature nodules may indicate that the triacylglycerides are stored for overwintering, as alfalfa nodules are known to be perennial. The oleosomes in root nodules of alfalfa do not appear to be directly associated with nitrogen fixation per se.Key words: oleosomes, root nodules, Medicago sativa, Rhizobium, pseudonodules.

Gene Expression of Medicago sativa Inoculated with Sinorhizobium meliloti as Modulated by the Xenobiotics Cadmium and Fluoranthene

2000 ◽  
Vol 55 (3-4) ◽  
pp. 222-232 ◽  
Author(s):  
Heike Neumann ◽  
Dietrich Werner
Keyword(s):  
Gene Expression ◽  
Arabidopsis Thaliana ◽  
Medicago Sativa ◽  
Sinorhizobium Meliloti ◽  
Rhizobium Meliloti ◽  
Root Nodules ◽  
Cadmium Concentration ◽  
Transcript Level ◽  
Northern Hybridization ◽  
Fresh Weight

Abstract Alfalfa plants (Medicago sativa cv. Europe) inoculated with Sinorhizobium meliloti 2011 (formerly Rhizobium meliloti, de Lajudie et al., 1994) were cultivated for 14 days under standardized growth conditions in mineral medium with addition of the heavy metal cadmium or the polycyclic aromatic hydrocarbon fluoranthene. These xenobiotics significantly reduced the numbers of root nodules before any visible damage to the plant could be detected. EC10. EC50, and EC90 (effective concentrations reducing nodulation, shoot and root fresh weight by 10, 50, or 90% compared to the control without pollutant) were calculated. EC50 for cadmium ranged from 5.8 jam (nodulation) to more than 20 μᴍ (root fresh weight). Testing fluoranthene resulted in an EC50 of 2.5 μg cm-2 for nodulation, and EC50 values of more than 35 μg cm-2 for shoot and root biomass production, indicating that the effect parameter nodulation is 10-fold more sensitive than shoot and root fresh weight. With m RNA differential display techniques the effects of both xenobiotics on gene expression in alfalfa root systems were studied. 37 differentially displayed transcripts were detected. Two of them, called DDMs1 and DDMs2, were confirmed by northern hybridization to be down-regulated in the presence of the xenobiotics. The expression of transcript DDMs1 was enhanced in alfalfa control plants inoculated with rhizobia, the transcript level was increased 2.5-3-fold compared to non-inoculated plants. This positive effect of nodulation was suppressed, partly by 35 μg cm-2 fluoranthene and totally by 20 μᴍ cadmium. The decrease in DDMsl transcription was highly affected by the cadmium concentration with an EC50 of 5.9 μᴍ . Compared to nodulation, almost identical EC10, EC50. and EC90 values were found for DDMsl expression. Sequence analysis of DDMsl revealed a significant overall homology (50% identity) to a hypothetical protein from Arabidopsis thaliana with high similarity to a copper transporting ATPase. High levels of transcript DDMs2 were observed in control plants with a 50% decrease in the xenobiotic-treated plants. DDM s2 gave a strong homology (82% identity) to the cytoplasmatic 60S ribosomal protein L18 from Arabidopsis thaliana.

Nodule morphogenesis: the early infection of Alfalfa (Medicago sativa) root hairs by Rhizobium meliloti

1989 ◽  
Vol 67 (10) ◽  
pp. 3108-3122 ◽  
Author(s):  
Susan M. Wood ◽  
William Newcomb
Keyword(s):  
Medicago Sativa ◽  
Root Hair ◽  
Rhizobium Meliloti ◽  
Root Hairs ◽  
Growth Period ◽  
Active Phase ◽  
Infection Threads ◽  
Thread Formation ◽  
Time Of Infection ◽  
Meliloti Strain

The growth and development of alfalfa (Medicago sativa) cv. Saranac root hairs and their infection by Rhizobium meliloti strain 102F51 was studied with Smith's interference contrast optics. Uninoculated root hairs grew and matured over a 10-h growth period. The nucleus migrated from a position opposite that of root-hair protrusion at initiation to the base of the root-hair protrusion, then into the growing root hair during the most active phase. When growth was nearly complete, the nucleus assumed a position near the base of the vacuolate root hair. If root hairs were inoculated during the first 2 h of growth after initiation, either "Shepherd's crooks" or root hairs deformed into a tight curl as the tip developed. Some of these Shepherd's rooks later demonstrated typical infection-thread formation. Root hairs that were inoculated between 4 and 6 h after root-hair initiation demonstrated branched growth, with the branch forming opposite the position of the nucleus at the time of infection. Infection threads occasionally formed in either the side branches or tip branches. Root hairs that were older than 6 h at the time of inoculation formed a variety of growth deformations, including ballooning, and elongate, spatulate, spiralling, or intertwined growth. Infections in this population of root hairs were rare.

Kinetics of first-cutting regrowth of alfalfa plants and nitrogenase activity in a controlled environment with and without added nitrate

1983 ◽  
Vol 61 (9) ◽  
pp. 2405-2409 ◽  
Author(s):  
F. D. H. Macdowall
Keyword(s):  
Dry Matter ◽  
Rhizobium Meliloti ◽  
Nitrogenase Activity ◽  
Radical Change ◽  
N Nutrition ◽  
Meliloti Strain ◽  
Medicago Sativa L ◽  
Reducing Activity ◽  
Acetylene Reducing Activity ◽  
Kinetics Of

Seedlings of Medicago sativa L. cv. Algonquin were grown in vermiculite and nodulated by Rhizobium meliloti strain 102F70 at two lower levels of N, until flowering when the tops were cut off to leave about 10% shoot stubble. Residual shoot dry matter immediately resumed first-order growth and maintained it throughout regrowth to second flowering. The rate constants of shoot regrowth were 34% lower (at 15 mM NO3−), 25% lower (at 1.5 mM NO3− symbiotically), or 220% higher (at zero NO3− symbiotically) than the values for 1 to 4-week-old seedlings, which indicated a radical change in physiology. Root dry matter resumed exponential growth after a 7-day recession and its recovery and yields were independent of N nutrition. The most pronounced minima occurred in the acetylene-reducing activity of nitrogenase, the kinetics of which paralleled root dry matter except that its redevelopment stopped after two-thirds of the regrowth time. The rate coefficient for the redevelopment of nitrogenase activity equalled that for its development during the seedling stage, which suggested unchanged limitations on that process until its redevelopment stopped.

Detection of Rhizobium meliloti cells in field soil and nodules by polymerase chain reaction

1995 ◽  
Vol 41 (9) ◽  
pp. 816-825 ◽  
Author(s):  
R. J. Watson ◽  
C. Haitas-Crockett ◽  
T. Martin ◽  
R. Heys
Keyword(s):  
Polymerase Chain Reaction ◽  
Rhizobium Meliloti ◽  
Genetically Engineered ◽  
Chain Reaction ◽  
Pcr Inhibitor ◽  
Vertical Location ◽  
Meliloti Strain ◽  
Identification Of Bacteria ◽  
Polymerase Chain ◽  
Genetically Engineered Microorganism

A genetically marked Rhizobium meliloti strain, R692, was prepared by insertion of a 1.7-kb DNA segment from Tn903 between the nifHDK and fixABC genes in the nod megaplasmid. This DNA was used as a marker, detectable by polymerase chain reaction (PCR), for the specific identification of bacteria in soil samples and alfalfa nodules. This detection technique was tested by applying different titres of the marked strain to field plots seeded with alfalfa. Samples of soil and nodules were assayed for the presence of the marker DNA fragment by PCR using primers specific to the marker sequence. The experiments revealed that the bacteria could be detected directly in soil containing about 103–104 bacteria/g, but greater sensitivity was prevented by potent PCR inhibitors present in the samples. The titre of the bacteria in the soil decreased rapidly after inoculation, dropping about 10-fold per week. Tests of vertical location of the bacteria in soil cores showed that the bacteria were initially dispersed to a depth of 18 cm, and subsequently retained viability in the top 2–8 cm. As few as 10 marked R. meliloti per gram of soil resulted in its establishment at detectable levels in nodules. Application of about 104–105 bacteria/g soil was sufficient to give the maximum number of nodules per plant and resulted in 70–90% occupancy by the marked strain. Limited movement of the inoculant was detected by analysis of nodules from plants adjacent to the sites where the bacteria were applied, probably by movement in water. The experiments demonstrated the advantages of PCR for the monitoring of marked microorganisms in the environment.Key words: genetically engineered microorganism, PCR inhibitor, nitrogen fixation, nif and fix genes, genetic marker.

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