Mycoplasma-like Organisms Associated with Proliferation and Big Bud of Solanum lycocarpum St. Hil.

Mycoplasma-like organisms (MLOs) were found in sieve tubes of Solanum lycocarpum affected with prolferation and big bud. The MLOs were graft-transmitted to tomato plants which developed similar symptoms. The association of MLOs with affected but not with symptomless S. lycocarpum and tomato suggests that the abnormality is induced by MLOs. The possible role of S. lycocarpum as a reservoir of a strain of the tomato big bud is discussed.     

Assessment of heavy metal accumulation in Anzali wetland,Iran, using a submerged aquatic plant,Ceratophyllum demersum

The objective of this study in 2009 was to examine whether levels of cadmium (Cd), copper (Cu), zinc (Zn), lead (Pb) and chromium (Cr) were higher in the leaves than in the stems of a submerged aquatic plant Ceratophyllum demersum in Anzali wetland. Cadmium, Pb and Cr concentrations were highest in the leaves. The mean concentrations of Cd and Cr in the leaves at all the sampling sites ranged between 0.94–1.26 μg g^?1 and 1.03–2.71 μg g^?1, respectively. Lead also had its highest concentrations in the leaves. The mean concentration of Pb in the leaves at all sampling sites ranged between 7.49–11.88 μg g^?1. Copper and Zn concentrations were highest in the stems. The mean concentrations of Cu and Zn in the stems at all sampling sites ranged between 10.79–17.91 μg g^?1 and 19.89–40.01 μg g^?1, respectively. Cadmium and Pb concentrations were higher in the leaves than in the stems, while Zn concentration was higher in the stems than in the leaves. Accumulation of Cu and Cr in the organs of C. demersum was in descending order of leaf ～ stem, since there was no significant difference between their mean concentrations in the leaves and stems.     

Maintenance of MLOs (Mycoplasma-like Organisms) on Populus alba Micropropagation

Populus alba plantlet micropropagation has been used for maintaining MLOs more than three years in samples collected from infected trees in Paris. Symptoms were observed on plantlets obtained from subcultures of the apical, middle and basal parts of the stems, and from the roots. Fluorescent microscopy failed to detect MLOs in the apical part of the plantlets and showed that they increased near the roots. Electron microscopy confirmed their presence. Some root sieve tubes were completely packed with MLOs. The sensitivity of the two methods used, subculturing and microscopy, for detection of MLOs is discussed. The symptoms remained for one year in plants regenerated from diseased plantlets and grown in the greenhouse. Then they started to disappear.     

PCR detection of MLOs in quick decline-affected pear trees in Italy

Polymerase chain reaction (PCR) amplification, using primers derived from the 16S rRNA gene, followed by restriction fragment length polymorphism (RFLP) analysis with Alu I restriction endonuclease was used to detect myc-oplasma-like organisms (MLOs) associated with pear decline. MLOs were consistently detected in pear trees that suddenly wilted and died within a few days during summer, as well as in pears of the same orchards with symptoms similar to the slow form of pear decline. In both cases the same RFLP pattern was obtained. Declining pear trees were 5 to 8-yr-old cvs Williams, Kaiser and Max Red Bartlett grafted on to Pyrus communis seedling rootstocks. All the orchards affected by quick decline had severe attacks of pear psyllid (Cacopsylla pyri) during the year this study was performed and during the previous year. The results showed the suitability of DNA amplification by the polymerase chain reaction for the detection of pear decline MLOs and established that MLOs can be detected in infected tissues of dead trees.     

Studies of the mycoplasma-like organism (MLO) in spinach leaves affected by the aster yellows disease

Summary Mycoplasma-like organisms (MLO) were found in the phloem of spinach (Spinacia oleracea L.) leaves infected with the agent of aster yellows disease by means of the leafhopperMacrosteles fascifrons Stål. The MLOs occurred mainly in mature sieve elements but were recorded in occasional phloem parenchyma cells as well. The MLO showed the typical features of this organism. The majority were ovoid or spherical, some were irregular in form or elongated. The larger bodies were commonly accompanied by small bodies which appeared to originate from the larger by budding. Profiles suggesting binary fission and filamentous forms containing ovoid condensations of cytoplasm were present. The bounding membrane showed the typical trilaminate structure, and DNA-like fibrils were discernible in those MLOs that had an electron lucent central region. In the denser bodies the fibrils were obscured. The MLO ribosomes were distinctly smaller than those in the host cytoplasm. The MLOs were degenerating in phloem cells that were disorganizing and collapsing in response to the infection. Structures in host cells that may be confused with MLO are described.This work was supported by the National Science Foundation grant GB-35228 to K. E and by Hatch and California Statewide Critical Applied Research Funds to the Departmem of Cell Physiology, University of California, Berkeley, California. The authors thank ProfessorJulius H.Freitag for providing the original strains of the aster yellows agent.     

First record of dill powdery mildew caused by Erysiphe heraclei DC in Egypt

In Egypt, powdery mildew was observed for the first time on dill plants, during annual disease surveys of March–May 2003 and 2005. Typical symptoms of powdery mildew of dill plant (Anethum graveolens L.) were observed in Gharbeia Governorate. Symptoms of powdery mildew became common on leaves, stems inflorescences and fruits as white irregular areas. These symptoms appeared at vegetative and early flowering stages then gradually increased through fruiting and pre-maturity stages. Samples of infected leaflets, stem, inflorescences and fruits were collected for examination by light and scanning electron microscope (SEM). Microscopic examination revealed that conidiophores were short, erect–69 × 6–10 μm in dimension, conidia were observed without conspicuous fibrosin bodies singly, elliposid to ovoid 25–33 × 10–16 μm in dimension, and the length to width ratio of conidia ranged from 1.7 to 2.0 and were produced singly. Cylindrical foot cells (22.0 × 8.0 μm) were followed by one or two shorter cells (12.5 × 7.5 μm). In spring, the sexual stage (cleistothecia) appeared on infected leaves and stems in spherical, gregarious measures 105–117 (111) × 100– 87.5 μm in diameter. Each cleistothecium contained (2–4) round to ovoid asci, 45–55 (50) × 45–25 (35) μm in dimension. The ascus contained (3–4) ellipsoid to ovoid ascospores, 20–17.5 × 15–10 (13.2) μm. Cleistothecia appendages are simple myceloid branched tips measuring 80–200 (140) μm in length and 3–5 (4) μm in diameter. Based on the observations of the morphology of its anamorph and teleomorph stages, the causal agent of dill powdery mildew was identified as Erysiphe heraclei which is reported for the first time in Egypt.     

CO2 enrichment and its relationship to bioconversion of cellulosic biomass of sweet potato (Ipomoea batatas L.) into fermentable sugars

Sweet potatoes (Ipomoea batatas L. (Lam) ‘Georgia-Jet’) were grown in open field plots and open top chambers at CO₂ concentrations of 354, 431, 506, and 659 μl liter⁻¹ for 90 days. The leaves and stems after the harvest were used as substrates for the production of fermentable sugars. Elevated CO₂ concentrations increased the cellulose content of stems, being most pronounced at 506 μl liter⁻¹. Hemicellulose content of leaves and stems as well as lignin content of stems decreased as a result of CO₂ enriched environment resulted in increased conversion of cellulose into fermentable sugars. The saccharification was greater in stems than in leaves. It was also found that chemical pretreatment of stems and leaves enhanced the enzymatic hydrolysis and the yields of glucose were higher than those from untreated stems and leaves.     

Mycoplasmalike Organisms (MLOs) Associated With the Witches' Broom Disease of Poplar

The witches' broom disease has been recently observed on poplars in Paris and its suburbs. The incidence of the disease appeared to be considerably high along main roads. The electron microscopic examination of 350 nm thick sieve tube sections revealed the presence of wall-less mycoplasmalike organisms (MLOs) in diseased samples of Populus alba var. nivea Wesm. They could not be found in their healthy counterparts.     

Association of the Decline of Nashi Pears with an MLO

Mycoplasma-like organisms (MLOs) were constantly detected by the DAPI technique and by restriction fragment length polymorphism (RFLP) analysis of PCR-amplified DNA in trees of Pyrus pyrifolia cvs Hosui and Kosui grafted on P. communis, seedlings rootstock with symptoms similar to the slow form of pear decline. These symptoms included upward curling of the leaves along the midrib. Leaves were abnormally thick and later turned reddish while major veins became swollen and brown. Trees with symptoms were usually 4–5 years old and were growing in the major pear areas of central Italy. The incidence of affected trees was particularly high in one orchard adjacent to a pear orchard strongly affected with the slow form of pear decline. In this case the distribution pattern of affected Nashi trees suggests that the causal agent was introduced from the adjacent pear orchard by an aerial vector. Although oriental pears are well-known hosts of the pear-decline agent when used as rootstocks of French cultivars, this is the first report of pear decline in P. pyrifolia varieties.     

LIGHT AND ELECTRON MICROSCOPY OF GRAZING BY POTERIOOCHROMONAS MALHAMENSIS (CHRYSOPHYCEAE) ON A RANGE OF PHYTOPLANKTON TAXA1

Grazing of fluorescent latex beads, bacteria, and various species of phytoplankton by Poterioochromonas malhamensis (Pringsheim) Peterfi (about 8.0 μm in diameter) was surveyed. The alga ingested fluorescent beads and various live or killed and nomnotile or motile organisms including bacteria, blue-green algae, green algae, diatoms, and chrysomonads. The size range of grazed prey was from 0.1 to 6.0 μm for latex beads and from 1.0 μm (bacteria) to about 21 μm (Carteria inverse) for organisms. As many as 17 latex beads (2.0 μm) or more than 10 Microcystis cells (5–6 μm) were ingested by a single P. malhamensis cell. Following such grazing, the cell increased in volume by up to about 30-fold. The range of cell volume of ingested prey was from 0.52 μm³ (bacteria) to about 3178 μm³(Carteria inversa). This study demonstrates for the first time that P. malhamensis is capable of grazing algae 2–3 times larger in diameter than its own cell and of grazing intact motile algae. Poterioochromonas malhamensis is an omnivorous grazer. Food vacuole formation and digestion processes were examined. The membrane that was derived from the plasma membrane and surrounded the prey disappeared sometime after ingestion. The food vacuole was then formed by successive fusion of numerous homogeneous vesicles accumulated around the prey. The prey was enclosed in a single membrane-bound food vacuole and then digested.     

CHARACTERIZATION OF STARCH GRAINS IN THE NONARTICULATED LATICIFER OF EUPHORBIA PULCHERRIMA (POINSETTIA)

Starch grain morphology in laticifer amyloplasts of Euphorbia pulcherrima Willd. (poinsettia) was examined for evidence of starch metabolism in vegetative and flowering plants. Laticifer starch grains in vegetative plants were rod shaped with lengths ranging from 3 to 60 μm. Average grain size was significantly larger in stems than leaves, and in older than younger tissues. Starch grain length frequency was unimodal and approximated a normal probability distribution in stems, but was skewed positively toward smaller grains in leaves. Frequency distributions were shifted toward larger grains in older tissues. Under short-day photoperiod (flowering) conditions, round starch grains formed in latex of stems, and the average length of rod-shaped grains decreased in latex of stems and leaves. Round grains did not occur in laticifers of leaves or bracts. Round starch grains often occurred in aggregates of two or more subunits. Changes in size and shape of latex starch grains indicate that amyloplasts in fully differentiated laticifers metabolize starch. Identification of metabolically active amyloplasts in differentiated laticifers suggests that the function of these organelles may involve starch mobilization under certain physiological conditions.     

Amplification of plasmid DNA to detect plant pathogenic mycoplasmalike organisms

The polymerase chain reaction (PCR) was employed to develop a specific assay for plant pathogenic mycoplasmalike organisms (MLOs). A cloned fragment of a plasmid from a severe strain of western aster yellows (AY)-MLO was sequenced to identify oligonucleotide primers for PCR. Amplified DNA fragments of the predicted size were obtained from DNA extracted from plants and insects infected with pear decline MLO, beet leafhopper-transmitted virescence agent, elm yellows MLO and several AY-MLO strains. No amplification occurred from healthy leafhopper or plant DNA. The PCR-based assay was over 500 times more sensitive than a _tilized_tion-based assay which _tilized a cloned AY plasmid fragment as a probe. With the PCR-based assay, MLOs could be detected using DNA samples of leafhoppers that were only crushed and boiled in buffer. Amplification of the target DNA was confirmed by digestion of the PCR product with Mbo I which yielded predicted sized fragments for all MLO strains except Bradford AY and eastern AY. Sequencing the PCR product from elm yellows and eastern AY-MLOs revealed greater than 90% homology, and the failure to restrict the PCR product with Mbo I was due to a single base change in the restriction endonuclease site. The ability of the assay to detect a wide range of MLOs with minimal sample preparation and high sensitivity will be useful in epidemiological studies of MLO-caused diseases.     

Antimicrobial Activity of Scabiosa arenaria Forssk. Extracts and Pure Compounds Using Bioguided Fractionation

The emergence of multidrug resistant pathogens threatened the clinical efficacy of many existing antibiotics. This situation has been recognized globally as a serious concern and justifies further research to discover antimicrobial agents from natural origins including plant extracts. The aim of our work was to evaluate the antimicrobial activities of Scabiosa arenaria Forssk . extracts and pure compounds using a bioguided fractionation, and try to explain some traditional use of this genus. The best antimicrobial activity‐guided fractionation was obtained by BuOH fractions of flowers, fruits and (stems and leaves) against Escherichia coli, Pseudomonas aeruginosa and Candida albicans with minimum inhibitory concentration (MIC) values from 0.0195 to 5 mg/ml. Escherichia coli was the most affected bug, thus the MIC of fruits BuOH extract showed the best anti‐Escherichia coli activity (MIC = 0.0195 mg/ml), followed by the (stems and leaves) and flowers BuOH extracts; MIC = 0.078 and 0.15 mg/ml, respectively. Furthermore, the subfractions obtained from these three mixed fractions showed also an important antimicrobial activity against the three microorganisms, with MIC values between 0.0195 and 0.312 mg/ml. The fractionation of the aerial part BuOH fraction led to the isolation of oleanolic acid ( 1 ) and luteolin 7‐O‐glucopyranoside ( 2 ) which are reported here for the first time from S. arenaria. Both compounds showed good antimicrobial activities with MIC values ranging from 170 to 683 μm and 86 to 347 μm , respectively. These results support the use of the Scabiosa genus to inhibit the growth of tested pathogenic bacteria and yeasts which may reduce illnesses associated with their exposure.     

Succession of microfungal flora on Rodgersia podophylla plants at the forest side of Cryptomeria plantation

Microfungal flora on aerial parts of a forest floor plant, Rodgersia podophylla, were studied at the forest side of a Japanese cedar plantation. From May to October, Nigrospora spp. were dominantly isolated from leaves, whereas Acremonium spp. and Fusarium spp. were dominant on stems, suggesting that the aerial part of the plants about 1 m height can offer two different habitats for these dominant fungi. In September and October, we could easily discern different types of tissue on the withering leaves, i.e., brown (necrotic lesion), yellowish (border tissue between brown and green areas), and green (healthy tissue). Nigrospora spp. and Pestalotiopsis spp. were continuously isolated on the brown area as well as on green and yellowish areas. Pathogenicity of N. sacchari and P. neglecta on potted plant leaves was confirmed by inoculation. From these, the fungi of these two genera seemed to have changed from quiescent to pathogenic with leaf senescence. Sporulations by fungi of the two genera were recognized on overwintered stems. These fungi may overwinter in stems that are slow to decompose, and seem to go over to the leaves in the following spring. Thus, they could be candidates for parasites that may play an important role in decomposition of the plant.     

Correlation between Light and Electron Microscopic Observations and Identification of Mycoplasmalike Organisms using Consecutive 350 nm Thick Sections

Serial sections of 350 nm thickness were used to make a correlation between electron and light microscopic observations. While thionin-acridine orange staining gave a positive result to detect abnormal sieve tubes of phyllody affected Phlox drummondii Hook, when observed under light microscope, the same cells revealed the presence of typical mycoplasmalike organisms (MLOs) in electron microscopic examination. Advantage of 350 nm thick sections in electron microscopy, and the utility of the technique in MLO detection have been discussed.     

Dispersal of Rhynchosporium secalis conidia from infected barley leaves or straw by simulated rain

Simulated rain (mean drop diameter c. 1 or 3 mm) was allowed to fall for 10 – 15 min on to barley leaves or straw infected by Rhynchosporium secalis (leaf blotch). The leaves were supported on a mesh through which run-off water drained and the straw was supported on a rigid surface on which run-off water collected. The numbers of R. secalis conidia and spore-carrying splash droplets collected by horizontal samplers (microscope slides and pieces of photographic film) decreased rapidly with increasing distance from and increasing height above the sources, with half-distances of 2 – 10 cm. Less than 10% of the spores or droplets reached heights of more than 30 cm. Incident drops 3 mm in diameter produced more spore-carrying droplets and dispersed more conidia than did 1 mm drops. The size category of splash droplets with the greatest proportion of the spore-carrying droplets dispersed by 3 mm drops was 200 – 400 μm, whether the source was infected barley leaves or barley straw. For leaves or straw the greatest proportions of spores were carried in droplets > 1000 μm in diameter. The mean diameter of spore-carrying droplets (478 μm) dispersed from free-draining leaves was less than that of droplets from straw plus run-off water (563 μm). However, the leaf source had more spores cm^-2 and the mean number of spores per droplet was greater (113 as opposed to 6·8) than for the straw source.     

Cytotoxic ent‐Kaurane Diterpenoids from Isodon nervosus

Four new ent‐kaurane diterpenoids, rabdonervosins G–J ( 1 – 4 , resp.), were isolated from the leaves and stems of Isodon nervosus. Their structures were elucidated by extensive spectroscopic analyses, including 1D‐, 2D‐NMR and HR mass spectra. Compound 2 showed potent cytotoxicity against the HepG2 and PC‐9/ZD cell lines with IC₅₀ values of 2.36 and 6.07 μM , respectively, and compound 3 exhibited cytotoxicity against the HepG2 and CNE2 cell lines with IC₅₀ values of 8.64 and 9.77 μM , respectively.     

Histopathology of Apple Proliferation in Malus Taxa and Hybrids of Different Susceptibility

Malus taxa and hybrids (“taxa”) grafted with M. pumila cv.‘Golden Delicious’differ significantly in their susceptibility to apple proliferation which is caused by a mycoplasma-like organism (MLO). These differences are correlated with the severity of anatomical aberrations and the numbers of MLOs in the phloem. The roots of declining trees of highly susceptible taxa with a mortality of more than 50 % are characterized by extensive phloem necrosis and the depletion of starch. MLOs are either not detectable or are present in low numbers, or the population appears degenerate when viewed by fluorescence microscopy. In comparable trees of a hybrid of M. sieboldii×M. pumila which shows a high recovery rate, both phloem necrosis and starch depletion are less pronounced, and the MLO numbers are low or the organisms are not detectable. Decline-tolerant taxa such as M. silvestris or M. pumila×M. baccata are little affected. Their phloem conditions and starch content do not differ significantly from that of healthy trees. However, the MLO titer is high. The histopathology of the scion cultivar of all groups examined is rather similar to that of the roots of the decline-tolerant taxa. Only in a late stage of decline, phloem necrosis increases while starch content and MLO numbers decrease in the scions grafted onto highly susceptible stockls.     

Chemical Composition and Antibacterial Activity of Essential Oils from Different Parts of Litsea cubeba

The composition of the essential oils obtained by hydrodistillation of different parts of Litsea cubeba, including roots, stems, leaves, alabastra (flower buds), flowers, and fruits, were investigated by GC (RI) and GC/MS. The antimicrobial activity of the oils was assessed with disc diffusion and microbroth dilution assays. The results showed large variations in the composition among the different oils. The major components in the oils from roots and fruits, from stems, leaves, and alabastra, and from flowers were citral B (neral), β‐phellandrene, and β‐terpinene, respectively. The inhibition zone (DD) and MIC values for the bacterial strains tested, which were all sensitive to the essential oil of L. cubeba, were in the range of 10.1–35.0 mm and 100–1000 μg/ml, respectively. Hence, the oils of the various parts showed moderate activity against the tested bacteria. This investigation showed that the antibacterial activity of L. cubeba was attributed to the essential oils, thus they can be a potential medicinal resource.     

CHLOROPLASTS OF MONOPLOID AND DIPLOID OENOTHERA HOOKERI

Comparison of leaves from diploid Oenothera hookeri plants with leaves of their monoploid derivatives revealed the following differences: the leaves of the diploids are almost twice as thick as those of the monoploids; the diploids have only one-fifth as many cells per unit leaf area as do the monoploids; leaf tissue of the diploids has an average of 36.3 chloroplasts per cell compared with 27.4 for the monoploids; chloroplasts of the diploids average 4.68 μm in length, those of the monoploids, 3.74 μm; chloroplasts from the diploids contain about seven times more chlorophyll than those of the monoploids and have an average of seven thylakoids per granum compared with an average of only three for the monoploids; the average chlorophyll a/b ratio for the diploid chloroplasts is 2.98 compared with 3.48 for the monoploids.