Comparative analyses of the complete genome sequences of Pierce's disease and citrus variegated chlorosis strains of Xylella fastidiosa

Xylella fastidiosa is a xylem-dwelling, insect-transmitted, gamma-proteobacterium that causes diseases in many plants, including grapevine, citrus, periwinkle, almond, oleander, and coffee. X. fastidiosa has an unusually broad host range, has an extensive geographical distribution throughout the American continent, and induces diverse disease phenotypes. Previous molecular analyses indicated three distinct groups of X.fastidiosa isolates that were expected to be genetically divergent. Here we report the genome sequence of X. fastidiosa (Temecula strain), isolated from a naturally infected grapevine with Pierce's disease (PD) in a wine-grape-growing region of California. Comparative analyses with a previously sequenced X.fastidiosa strain responsible for citrus variegated chlorosis (CVC) revealed that 98% of the PD X.fastidiosa Temecula genes are shared with the CVC X. fastidiosa strain 9a5c genes. Furthermore, the average amino acid identity of the open reading frames in the strains is 95.7%. Genomic differences are limited to phage-associated chromosomal rearrangements and deletions that also account for the strain-specific genes present in each genome. Genomic islands, one in each genome, were identified, and their presence in other X.fastidiosa strains was analyzed. We conclude that these two organisms have identical metabolic functions and are likely to use a common set of genes in plant colonization and pathogenesis, permitting convergence of functional genomic strategies.

Structure of a glycoglucuronomannan from the low-viscosity gum of Vochysia lehmannii

The polysaccharide (VSP) from the gum exudate of quaruba (Vochysia lehmannii) had two components of almost identical M. centred at 24,800, as shown by HSPEC-MALLS. The presence of aggregates was shown since carboxy-reduction gave VSP-RED, which contained low molecular weight components with M-w 19,000 > 5800 and polydispersity ratios dn/dc 0.160 and 0.149, respectively. VSP formed low viscosity aqueous solutions and acid hydrolysis gave Man (30%), Ara (16%), Gal (10%), and Glc (14%). The latter arose partly from GlcA (30%). Methylation analysis revealed mainly neutral units of 2-O- (60%) and 2,3-di-O-substituted Manp (5%), and those of nomeducing ends (8%), 2-O- (3%), and 4-O-substituted Arap and/or 5-O-substituted Araf units (6%). VSP-RED contained Glc (45%), Man (35%), and Ara (13%) and methylation analysis indicated mainly 4-O-substituted Glcp (31%) and 2-O- (51%) and 2,3-di-O-substituted Manp units (5%). A predominant alternating structure for VSP was shown by its C-13 NMR spectrum, which contained 10 main signals and a small one of C-6 of GlcpA. This was confirmed by formation, on partial hydrolysis of VSP, of a tetrasaccharide, which was characterised by NMR spectroscopy and ESI-MS as beta-GlcpA-(1 --> 2)-alpha-Manp-(1 --> 4)-beta-GlcpA-(1 --> 2)-Man, which arose from the main chain, thus confirming VSP to be a glycoglucuronomannan. (C) 2004 Elsevier Ltd. All rights reserved.

Recombinant expression and characterization of a Xylella fastidiosa cysteine protease differentially expressed in a nonpathogenic strain

Xylella fastidiosa is a xylem-limited, Gram-negative bacterium responsible for citrus variegated chlorosis (CVC) in sweet oranges. In the present study, we present the recombinant expression, purification and characterization of an X. fastidiosa cysteine protease (dubbed Xylellain). The recombinant Xylellain ((HIS)Xylellain) was able to hydrolyze carbobenzoxy-Phe-Arg-7-amido-4-methylcoumarin (Z-FR-MCA) and carbobenzoxy-Arg-Arg-7-amido-4-methylcoumarin (Z-RR-MCA) with similar catalytic efficiencies, suggesting that this enzyme presents substrate specificity requirements similar to cathepsin B. The immunization of mice with (HIS)Xylellain provided us with antibodies, which recognized a protein of c. 31 kDa in the X. fastidiosa pathogenic strains 9a5c, and X. fastidiosa isolated from coffee plants. However, these antibodies recognized no protein in the nonpathogenic X. fastidiosa J1a12, suggesting the absence or low expression of this protein in the strain. These findings enabled us to identify Xylellain as a putative target for combating CVC and other diseases caused by X. fastidiosa strains.

Shelf-life of chicken loaves

This research was designed to study the elaboration of chicken loaf with ground meat. The products were elaborated in laboratory scale utilizing broiler meat bought at the market. The products were conditioned in vacuum-packages and stored for 133 days at 5 and 10 degrees C. Chemical, sensorial and microbiological analyses of the elaborated products were performed. The storage temperature affected the total bacterial counts, pH, characteristic aroma and rancid aroma. In both treatments (storage temperature: 5 and 10 degrees C), depletion of nitrites, reaching 24 ppm in 133 days of storage, and exudate formation after 77 days were observed. The storage of chicken leaves, vacuum-packaged, at 5 degrees C for a maximum of 77 days is recommended.

Kinetics of zinc uptake and anatomy of roots and leaves of coffee trees as affected by zinc nutrition

Zinc (Zn) uptake kinetics and root and leaf anatomy were studied in coffee trees grown in nutrient solutions with or without Zn. Leaves and roots were sampled and cuts were made in the medium part of the leaves and in root tips and observed under an optical microscope. Plants grown without Zn showed an increase in root and in root stele diameter. There was also an increase in epidermis thickness and in the cross-sectional area of the cortex and stele due to Zn deficiency, but the diameter of xylem vessels was decreased. An increase in root cortex and stele diameter provided for an increased surface for nutrient uptake. Accordingly, C(min) was decreased from 13.8 to 3.4 mu mol L(-1) and V(max) increased from 0.50 to 2.1 mu mol cm(-2) h(-1) .

Boron deficiency inhibits petiole and peduncle cell development and reduces growth of cotton

The effect of boron (B) on cotton growth and fruit shedding may be due not only to physiological or biochemical effects, but also to vascular tissue malformation. This experiment investigated petiole and floral peduncle anatomical alterations and growth of cotton supplied with deficient and sufficient B in nutrient solution. Cotton (Gossypium hirsutum cv. 'Delta Opal') plants were grown in solutions containing 0, 1.5, 3.0, 4.5, and 6.0 mu mol L-1 of B from 22 to 36 d after plant emergence (DAPE). From 36 to 51 DAPE, B was omitted from the nutrient solution. Petioles from young leaves and floral bud peduncles (first position of the first sympodial) were sampled and the cross-section anatomy observed under an optical microscope. The number of vascular bundles of the petiole was decreased in B-deficient plants and the xylem was disorganized. Phloem elements in the peduncle vascular cylinder of B-deficient plants did not show clear differentiation. The few xylem elements that were formed were also disorganized. Modifications caused by B deficiency may have impaired B and photosynthate translocation into new cotton growth. Boron accumulation in the shoot of B-deficient plants suggested that there was some B translocation within the plant. It could be inferred that cotton growth would be impaired by the decrease in carbohydrate translocation rather than by B deficiency in the tissue alone.

Propagação de sapotizeiro (Manilkara zapota L.) por estaquia

Two sapoti tree accesses (Manilkara zapota), native plants of south of Mexico and of America Central, belonging to the Collection of Fruitful Native and Exotic of UNESP/FCAV, Jaboticabal Campus, was studied with relationship to the cutting propagation. The first test consisted of fast immersion of cutting base semi-herbaceous foliage of selected accesses FCAV-III and FCAV-VII. The ttreatments were alcoholic solution (50%) of indolbutiric acid (IBA) in concentrations of 0, 1,000, 3,000, 5,000 and 7,000 mgL(-1). and the cuttings preparing with and without latex exudate washing for a period of 14 hours in recipient with pure water. This trial was set in 5 x 2 x 2 factorial scheme (IBA concentrations x accesses x cutting preparation) The second test consisted the herbaceous foliage immersion of FCAV-III access in IBA concentrations above. Both experiments were distributed in randomized design with four replications and 10 cuttings each. All cuttings were collected in spring and prepared with about 12 cm of length, planted in medium texture vermiculite and carried under conditions intermittent mist spray flashing in chamber with 50% light. The two accesses tested not rooted and treatments were not able to induce the roots emission.

Seasonal presence of acicular calcium oxalate crystals in the cambial zone of Citharexylum myrianthum (Verbenaceae)

This study focuses on the seasonal presence of acicular crystals in the cambial zone of Citharexylum myrianthum Chain. (Verbenaceae). Specimens collected in different months from 1996 to 2000 were examined for the abundance of acicular crystals in the cambium. This information was correlated with the phenology of the species and the climate of the region. Acicular calcium oxalate crystals were found in cambial fusiform and ray cell initials, as well as in their daughter cells. An abundance of crystals was observed during periods of water deficit and leaf fall (July). Fewer crystals were found in the beginning of the wet season and bud swelling (September). When trees were flowering and the soil was wet (November and December), acicular crystals were rarely observed. During this period, acicular crystals were found in differentiating phloem and xylem parenchyma cells, in fully differentiated phloem cells, but not in fully differentiated xylem cells.

Lactate in Bitches with Pyometra

Lactate is a compound produced by the anaerobic metabolism of glucose, and hyperlactataemia occurs when the rate of production of lactate exceeds the rate of elimination. This occurs in situations of hypoxia and tissue hypoperfusion. Lactate has been considered a useful prognostic indicator in critically ill patients. Pyometra is a disease of adult female dogs characterized by inflammation of the uterus with an accumulation of exudate, which occurs during the luteal phase. It is one of the most common diseases that occur in the genital tract of female dogs. A total of 31 dogs were diagnosed with pyometra. The diagnosis was confirmed at ultrasonography. of the 31 dogs, 25 females had open cervix pyometra and six had closed cervix pyometra. Plasma lactate concentrations were determined by an enzymatic colorimetric method. The average concentration (+/- SD) of plasma lactate in all 31 bitches with pyometra was 3.55 +/- 0.46 mm. Healthy dogs had plasma lactate concentrations between 0.3 and 2.5 mm (mean +/- SD). Concentrations ranged from 0.8 to 2.9 mm when plasma lactate was measured with a portable device and 0.42.6 mm with the blood gas analyser. Even though plasma lactate values vary between several studies and equipment used to measure concentrations, our results for dogs with pyometra are higher indicating hyperlactataemia (Thorneloe et al. , Can Vet J 48, 283288). Plasma lactate in dogs with closed cervix pyometra was mean +/- SD and in dogs with open cervix pyometra, it was mean +/- SD. The plasma lactate concentration in dogs with pyometra was higher than in healthy bitches, and there was no influence of patency of the cervix on the concentration of plasma lactate concentrations. Plasma lactate concentrations were similar for animals with open and closed pyometra (3.54 +/- 0.52 to 3.64 +/- 1.03 mm).

Development of Intra- and Interxylary Secondary Phloem in Coccinia indica (Cucurbitaceae)

Stern anatomy and the development of intraxylary phloem were investigated in six to eight years old Coccinia indica L. (Cucurbitaceae). Secondary growth in the stems was achieved by the normal cambial activity. In the innermost part of the thicker stems, xylem parenchyma and pith cells dedifferentiated into meristematic cells at several points. In some of the wider rays, ray cells dedifferentiate and produce secondary xylem and phloem with different orientations and sometimes a complete bicollateral vascular bundle. The inner cambial segments of the bicollateral vascular bundle (of primary growth) maintained radial arrangement even in the mature stems but in most places the cambia were either inactive or showed very few cell divisions. Concomitant with the obliteration and collapse of inner phloem (of bicollateral vascular bundles), parenchyma cells encircling the phloem became meristematic forming a circular sheath of internal cambia. These internal cambia produce only intraxylary secondary phloem centripetally and do not produce any secondary xylem. In the stem, secondary xylem consisted mainly of axial parenchyma, small strands of thick-walled xylem derivatives, i.e. vessel elements and fibres embedded in parenchymatous ground mass, wide and tall rays along with exceptionally wide vessels characteristic of lianas. In thick stems, the axial parenchyma de-differentiated into meristem, which later re-differentiated into interxylary phloem. Fibre dimorphism and pseudo-vestured pits in the vessels are also reported.

alpha-Hydroxynitrile lyase protein from Xylella fastidiosa: Cloning, expression, and characterization

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

A putative twin-arginine translocation system in the phytopathogenic bacterium Xylella fastidiosa

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Copper resistance of biofilm cells of the plant pathogen Xylella fastidiosa

Xylella fastidiosa is a phytopathogen that causes diseases in different plant species. The development of disease symptoms is associated to the blockage of the xylem vessels caused by biofilm formation. In this study, we evaluated the sensitivity of biofilm and planktonic cells to copper, one of the most important antimicrobial agents used in agriculture. We measured the exopolysaccharides (EPS) content in biofilm and planktonic cells and used real-time reverse transcription polymerase chain reaction to evaluate the expression of the genes encoding proteins involved in cation/multidrug extrusion (acrA/B, mexE/czcA, and metI) and others associated with different copper resistance mechanisms (copB, cutA1, cutA2, and cutC) in the X. fastidiosa biofilm formed in two different media. We confirmed that biofilms are less susceptible to copper than planktonic cells. The amount of EPS seems to be directly related to the resistance and it varies according to the media where the cells are grown. The same was observed for gene expression. Nevertheless, some genes seem to have a greater importance in biofilm cells resistance to copper. Our results suggest a synergistic effect between diffusion barriers and other mechanisms associated with bacterial resistance in this phytopathogen. These mechanisms are important for a bacterium that is constantly under stress conditions in the host.

Development of intraxylary phloem and internal cambium in Ipomoea hederifolia (Convolvulaceae)

V. S. PATIL (Department of Botany, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara-390002 India), K. S. RAO (BRD School of Bioscieces, S. P. University, Vallabh Vidyanagar, India), and K. S. RAJPUT (Department of Botany, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara-390002 India). Development of intraxylary phloem and internal cambium in Ipomoea hederifolia (Convolvulaceae). J. Torrey Bot. Soc. 136: 423-432. 2009-In Ipomoea hederifolia L. (Convolvulaceae), internal/intraxylary phloem originated as isolated strands from the procambially derived cells after the formation of protoxylem and protophloem. Bands of internal phloem were apparent in the sixth internode after the development of metacambium. In the relatively thick stems several small arcs/segments of internal cambium ensues from the parenchyma cells between the protoxylem and internal protophloem. Though all the segments were active, some of them (two of them located opposite to each other) were relatively more active. Bidirectional differentiation of these segments gave rise to secondary xylem centrifugally and secondary phloem centripetally, resulting inverted vascular bundles. Rest of the internal cambium segments were unidirectional and formed only secondary phloem centripetally. Like external vascular cambium, the internal cambium was non-storied. Structurally, secondary xylem and phloem was composed of axial and radial system in which rays were mostly uni- to biseriate. Secondary xylem produced by the internal cambium was more or less similar to the xylem formed by the external successive cambia. Secondary phloem produced by the internal cambium was composed of sieve tubes, companion cells, axial and ray parenchyma cells. Simple sieve plates of internal phloem were mostly arranged on transverse end walls in contrast to compound and obliquely placed sieve plates of external phloem formed by the successive cambia.

Development of successive cambia and structure of wood in Gallesia integrifolia (Spreng.) Harms (Phytolaccaceae)

Stem diameter in Gallesia integrifolia (Spreng.) Harms (Phytolaccaceae) increases by forming concentric rings of xylem alternating with phloem, which show frequent anastomoses. After a period of primary growth and the formation of first (normal) ring of vascular cambium, further successive rings are initiated outside this cambium. The second ring of cambium originates from the pericycle parenchyma located between the proto-phloem, and the pericycle fibres. Each cambium produces centripetally secondary xylem and centrifugally secondary phloem. Differentiation of xylem precedes that of phloem and the first elements formed are always xylem fibres. Structurally, the vascular cylinder is composed by successive rings of secondary xylem and phloem. These rings are separated by wide bands of conjunctive parenchyma tissue. Presence of collateral vascular bundles with irregular orientation is observed in the region of anastomoses of two or more bands of conjunctive tissue. These bundles are surrounded by isodiametric, lignified and thick-walled cells. In some of the cambial rings, occurrence of polycentric rays was also noticed; these rays are tall, and characterized by the presence of meristematic regions that differentiated into thick-walled elements of secondary xylem. Origin and development of the successive cambia and the structure of xylem are discussed.