Complete Genome Sequence of Mycoplasma suis and Insights into Its Biology and Adaption to an Erythrocyte Niche

Mycoplasma suis, the causative agent of porcine infectious anemia, has never been cultured in vitro and mechanisms by which it causes disease are poorly understood. Thus, the objective herein was to use whole genome sequencing and analysis of M. suis to define pathogenicity mechanisms and biochemical pathways. M. suis was harvested from the blood of an experimentally infected pig. Following DNA extraction and construction of a paired end library, whole-genome sequencing was performed using GS-FLX (454) and Titanium chemistry. Reads on paired-end constructs were assembled using GS De Novo Assembler and gaps closed by primer walking; assembly was validated by PFGE. Glimmer and Manatee Annotation Engine were used to predict and annotate protein-coding sequences (CDS). The M. suis genome consists of a single, 742,431 bp chromosome with low G+C content of 31.1%. A total of 844 CDS, 3 single copies, unlinked rRNA genes and 32 tRNAs were identified. Gene homologies and GC skew graph show that M. suis has a typical Mollicutes oriC. The predicted metabolic pathway is concise, showing evidence of adaptation to blood environment. M. suis is a glycolytic species, obtaining energy through sugars fermentation and ATP-synthase. The pentose-phosphate pathway, metabolism of cofactors and vitamins, pyruvate dehydrogenase and NAD(+) kinase are missing. Thus, ribose, NADH, NADPH and coenzyme A are possibly essential for its growth. M. suis can generate purines from hypoxanthine, which is secreted by RBCs, and cytidine nucleotides from uracil. Toxins orthologs were not identified. We suggest that M. suis may cause disease by scavenging and competing for host nutrients, leading to decreased life-span of RBCs. In summary, genome analysis shows that M. suis is dependent on host cell metabolism and this characteristic is likely to be linked to its pathogenicity. The prediction of essential nutrients will aid the development of in vitro cultivation systems.

Leishmania-Specific Surface Antigens Show Sub-Genus Sequence Variation and Immune Recognition

Background: A family of hydrophilic acylated surface (HASP) proteins, containing extensive and variant amino acid repeats, is expressed at the plasma membrane in infective extracellular (metacyclic) and intracellular (amastigote) stages of Old World Leishmania species. While HASPs are antigenic in the host and can induce protective immune responses, the biological functions of these Leishmania-specific proteins remain unresolved. Previous genome analysis has suggested that parasites of the sub-genus Leishmania (Viannia) have lost HASP genes from their genomes. Methods/Principal Findings: We have used molecular and cellular methods to analyse HASP expression in New World Leishmania mexicana complex species and show that, unlike in L. major, these proteins are expressed predominantly following differentiation into amastigotes within macrophages. Further genome analysis has revealed that the L. (Viannia) species, L. (V.) braziliensis, does express HASP-like proteins of low amino acid similarity but with similar biochemical characteristics, from genes present on a region of chromosome 23 that is syntenic with the HASP/SHERP locus in Old World Leishmania species and the L. (L.) mexicana complex. A related gene is also present in Leptomonas seymouri and this may represent the ancestral copy of these Leishmania-genus specific sequences. The L. braziliensis HASP-like proteins (named the orthologous (o) HASPs) are predominantly expressed on the plasma membrane in amastigotes and are recognised by immune sera taken from 4 out of 6 leishmaniasis patients tested in an endemic region of Brazil. Analysis of the repetitive domains of the oHASPs has shown considerable genetic variation in parasite isolates taken from the same patients, suggesting that antigenic change may play a role in immune recognition of this protein family. Conclusions/Significance: These findings confirm that antigenic hydrophilic acylated proteins are expressed from genes in the same chromosomal region in species across the genus Leishmania. These proteins are surface-exposed on amastigotes (although L. (L.) major parasites also express HASPB on the metacyclic plasma membrane). The central repetitive domains of the HASPs are highly variant in their amino acid sequences, both within and between species, consistent with a role in immune recognition in the host.

SOME PROPERTIES OF THE MULTIPLICITY SEQUENCE FOR ARBITRARY IDEALS

In this work we prove that the Achilles-Manaresi multiplicity sequence, like the classical Hilbert-Samuel multiplicity, is additive with respect to the exact sequence of modules. We also prove the associativity formula for his mulitplicity sequence. As a consequence, we give new proofs for two results already known. First, the Achilles-Manaresi multiplicity sequence is an invariant up to reduction, a result first proved by Ciuperca. Second, I subset of J is a reduction of (J,M) if and only if c(0)(I(p), M(p)) = c(0)(J(p), M(p)) for all p is an element of Spec(A), a result first proved by Flenner and Manaresi.

Sequence-specific electrochemical detection of Alicyclobacillus acidoterrestris DNA using electroconductive polymer-modified fluorine tin oxide electrodes

This study outlines the quantification of low levels of Alicyclobacillus acidoterrestris in pure cultures, since this bacterium is not inactivated by pasteurization and may remain in industrialized foods and beverages. Electroconductive polymer-modified fluorine tin oxide (FTO) electrodes and multiple nanoparticle labels were used for biosensing. The detection of A. acidoterrestris in pure cultures was performed by reverse transcription polymerase chain reaction (RT-PCR) and the sensitivity was further increased by asymmetric nested RT-PCR using electrochemical detection for quantification of the amplicon. The quantification of nested RT-PCR products by Ag/Au-based electrochemical detection was able to detect 2 colony forming units per mL (CFU mL(-1)) of spores in pure culture and low detection and quantification limits (7.07 and 23.6 nM, respectively) were obtained for the target A. acidoterrestris on the electrochemical detection bioassay.

Highly Toxic Microcystis aeruginosa Strain, Isolated from So Paulo-Brazil, Produce Hepatotoxins and Paralytic Shellfish Poison Neurotoxins

While evaluating several laboratory-cultured cyanobacteria strains for the presence of paralytic shellfish poison neurotoxins, the hydrophilic extract of Microcystis aeruginosa strain SPC777-isolated from Billings`s reservoir, So Paulo, Brazil-was found to exhibit lethal neurotoxic effect in mouse bioassay. The in vivo test showed symptoms that unambiguously were those produced by PSP. In order to identify the presence of neurotoxins, cells were lyophilized, and the extracts were analyzed by HPLC-FLD and HPLC-MS. HPLC-FLD analysis revealed four main Gonyautoxins: GTX4(47.6%), GTX2(29.5%), GTX1(21.9%), and GTX3(1.0%). HPLC-MS analysis, on other hand, confirmed both epimers, with positive Zwitterions M(+) 395.9 m/z for GTX3/GTX2 and M(+) 411 m/z for GTX4/GTX1 epimers. The hepatotoxins (Microcystins) were also evaluated by ELISA and HPLC-MS analyses. Positive immunoreaction was observed by ELISA assay. Alongside, the HPLC-MS analyses revealed the presence of [l-ser(7)] MCYST-RR. The N-methyltransferase (NMT) domain of the microcystin synthetase gene mcyA was chosen as the target sequence to detect the presence of the mcy gene cluster. PCR amplification of the NMT domain, using the genomic DNA of the SPC777 strain and the MSF/MSR primer set, resulted in the expected 1,369 bp product. The phylogenetic analyses grouped the NMT sequence with the NMT sequences of other known Microcystis with high bootstrap support. The taxonomical position of M. aeruginosa SPC777 was confirmed by a detailed morphological description and a phylogenetic analysis of 16S rRNA gene sequence. Therefore, co-production of PSP neurotoxins and microcystins by an isolated M. aeruginosa strain is hereby reported for the first time.

Nasopharyngeal Intubation in Robin Sequence: Technique and Management

Objective: To provide a detailed description of the nasopharyngeal intubation (NPI) technique and photographs, which should be helpful for those who may need to perform it for treating the airway obstruction in Robin sequence. Design: To describe and illustrate the NPI technique and the necessary considerations for its application. Setting: Hospital de Reabilitacao de Anomalias Craniofacial of University of Sao Paulo, Brazil. Result: The NPI procedure involves the use of a whitish, Portex, number 3.0 or 3.5, silicone tube, introduced 8 cm deep into the infant`s nostril and fixed with Micropore tape. The tube is to be removed at least twice a day for proper hygiene (with running water, detergent, and swabs) and should be changed every 7 days. This procedure is taught to the children`s parents or caretakers by the nurse during hospitalization. Conclusion: The technique is so simple that it can be performed by the parents themselves, allowing continuation of the treatment at home.

Number of options in a movement sequence affects learners' behavior in a self-controlled practice condition

Self controlling practice implies a process of decision making which suggests that the options in a self controlled practice condition could affect learners The number of task components with no fixed position in a movement sequence may affect the (Nay learners self control their practice A 200 cm coincident timing track with 90 light emitting diodes (LEDs)-the first and the last LEDs being the warning and the target lights respectively was set so that the apparent speed of the light along the track was 1 33 m/sec Participants were required to touch six sensors sequentially the last one coincidently with the lighting of the tar get light (timing task) Group 1 (n=55) had only one constraint and were instructed to touch the sensors in any order except for the last sensor which had to be the one positioned close to the target light Group 2 (n=53) had three constraints the first two and the last sensor to be touched Both groups practiced the task until timing error was less than 30 msec on three consecutive trials There were no statistically significant differences between groups in the number of trials needed to reach the performance criterion but (a) participants in Group 2 created fewer sequences corn pared to Group 1 and (b) were more likely to use the same sequence throughout the learning process The number of options for a movement sequence affected the way learners self-controlled their practice but had no effect on the amount of practice to reach criterion performance.

The Policy Iteration Algorithm for Average Continuous Control of Piecewise Deterministic Markov Processes

The main goal of this paper is to apply the so-called policy iteration algorithm (PIA) for the long run average continuous control problem of piecewise deterministic Markov processes (PDMP`s) taking values in a general Borel space and with compact action space depending on the state variable. In order to do that we first derive some important properties for a pseudo-Poisson equation associated to the problem. In the sequence it is shown that the convergence of the PIA to a solution satisfying the optimality equation holds under some classical hypotheses and that this optimal solution yields to an optimal control strategy for the average control problem for the continuous-time PDMP in a feedback form.

Characterization of electrical penetration graphs of Bucephalogonia xanthophis, a vector of Xylella fastidiosa in citrus

The sharpshooter Bucephalogonia xanthophis (Berg) (Homoptera: Cicadellidae) is a vector of the xylem-limited bacterium, Xylella fastidiosa (Wells, Raju, Hung, Weisburg, Mandelco-Paul, and Brenner), which causes citrus variegated chlorosis. Despite the importance of citrus variegated chlorosis, the probing behavior of vectors on citrus and its implications for transmission of X. fastidiosa have not been studied. Here we studied electrical penetration graph (EPG-DC system) waveforms produced by B. xanthophis on Citrus sinensis (L.) Osbeck (Rutaceae), and their relationships with stylet activities and xylem ingestion. Electrical penetration graph waveforms were described based on amplitude, frequency, voltage level, and electrical origin of the observed traces during stylet penetration on plant tissues. The main waveforms were correlated with histological observations of salivary sheaths in plant tissues and excretion analysis, in order to determine stylet activities and their precise position. Six waveforms and associated activities are described: (S) secretion of salivary sheath and intracellular stylet pathway, (R) resting during stylet pathway, (Xc) contact of stylets with xylem vessels, (Xi) active xylem ingestion, (N) interruption within the xylem phase (during Xc or Xi), and (W) withdrawal of stylet from the plant. The sharpshooter spent 91.8% of its probing time with its stylet in the xylem, where the main activity was ingestion (Xi: 97.5%). During a probe, the most likely sequence of events is secretion of salivary sheath and pathway (S) through epidermal and parenchyma cells (all individuals), followed by contact with xylem (Xc) (67.6% of all individuals) and ingestion (Xi) (88.3% of those that exhibit waveform Xc). The mean time to contact the xylem (Xc) and initiate ingestion (Xi) after onset of the first probe was 27.8 and 34.2 min, respectively. However, sustained xylem ingestion (Xi > 5 min) was established after 39.8 min, on average. This information is basic for future studies on the transmission mechanisms of X. fastidiosa and in order to establish control strategies aimed at interfering with this process.

MACRONUTRIENTS UPTAKE BY POTATO (Solanum tuberosum L.) IN SEED-TUBER PRODUCTION

In greenhouse potato cultivation, mineral nutrition is one of the main factors contributing to high yields and better product quality. Knowledge about the amount of nutrients accumulated in the plants at each growing phase provides important information that helps the establishment of a more balanced fertilizer application. The objective of this research was to determine the time course of macronutrients uptake and accumulation in potato plants for seed-tuber production, grown in nutrient solution. The experiment was carried out in a greenhouse, using in vitro material from the pre-basic category of the `Atlantic` variety. The plants were collected weekly from 14 days after transplanting (DAT) until 70 DAT The experimental design was a completely randomized block with 9 treatments to sampling times and four replicates. The highest nutrient requirement in the plant shoot occurred at the periods between 28 and 56 DAT while in the tubers it was after 49 DAT The maximum accumulation sequence of macronutrients was K > N > S > Ca > P > Mg.

Biogeochemical cycles of nutrients in tropical Eucalyptus plantations Main features shown by intensive monitoring in Congo and Brazil

The sustainability of fast-growing tropical Eucalyptus plantations is of concern in a context of rising fertilizer costs, since large amounts of nutrients are removed with biomass every 6-7 years from highly weathered soils. A better understanding of the dynamics of tree requirements is required to match fertilization regimes to the availability of each nutrient in the soil. The nutrition of Eucalyptus plantations has been intensively investigated and many studies have focused on specific fluxes in the biogeochemical cycles of nutrients. However, studies dealing with complete cycles are scarce for the Tropics. The objective of this paper was to compare these cycles for Eucalyptus plantations in Congo and Brazil, with contrasting climates, soil properties, and management practices. The main features were similar in the two situations. Most nutrient fluxes were driven by crown establishment the two first years after planting and total biomass production thereafter. These forests were characterized by huge nutrient requirements: 155, 10, 52, 55 and 23 kg ha(-1) of N, P, K, Ca and Mg the first year after planting at the Brazilian study site, respectively. High growth rates the first months after planting were essential to take advantage of the large amounts of nutrients released into the soil solutions by organic matter mineralization after harvesting. This study highlighted the predominant role of biological and biochemical cycles over the geochemical cycle of nutrients in tropical Eucalyptus plantations and indicated the prime importance of carefully managing organic matter in these soils. Limited nutrient losses through deep drainage after clear-cutting in the sandy soils of the two study sites showed the remarkable efficiency of Eucalyptus trees in keeping limited nutrient pools within the ecosystem, even after major disturbances. Nutrient input-output budgets suggested that Eucalyptus plantations take advantage of soil fertility inherited from previous land uses and that long-term sustainability will require an increase in the inputs of certain nutrients. (C) 2009 Elsevier B.V. All rights reserved.

Purification and Characterization of a Trypsin Inhibitor from Plathymenia foliolosa Seeds

A novel trypsin inhibitor (PFTI) was isolated from Plathymenia foliolosa (Benth.) seeds by gel filtration chromatography on a Sephadex G-100, DEAE-Sepharose, and trypsin-Sepharose columns. By SDS-PAGE, PFTI yielded a single band with a M(r) of 19 kDa. PFTI inhibited bovine trypsin and bovine chymotrypsin with equilibrium dissociation constants (K(i)) of 4 x 10(-8) and 1.4 x 10(-6) M, respectively. PFTI retained more than 50% of activity at up to 50 degrees C for 30 min, but there were 80 and 100% losses of activity at 60 and 70 degrees C, respectively. DTT affected the activity or stability of PFTI. The N-terminal amino acid sequence of PFTI showed a high degree of homology with various members of the Kunitz family of inhibitors. Anagasta kuehniella is found worldwide; this insect attacks stored grains and products of rice, oat, rye, corn, and wheat. The velvet bean caterpillar (Anticarsia gemmatalis) is considered the main defoliator pest of soybean in Brazil. Diatraea saccharalis, the sugar cane borer, is the major pest of sugar cane crops, and its caterpillar-feeding behavior, inside the stems, hampers control. PFTI showed significant inhibitory activity against trypsin-like proteases present in the larval midguts on A. kuehniella and D. saccharalis and could suppress the growth of larvae.

Surface Liming and Zinc Availability in a Long-Term Experiment under No-Till System

No-till (NT) system with crop rotation is one of the most effective strategies to improve agricultural sustainability in tropical and subtropical regions. To control soil acidity in NT, lime is broadcast on the surface without incorporation. The increase in soil pH due to surface liming may decrease zinc (Zn) availability and its uptake by crops. A field experiment was performed in Parana State, Brazil, on a loamy, kaolinitic, thermic Typic Hapludox to evaluate Zn bioavailability in a NT system after surface liming and re-liming. Dolomitic lime was surface applied on the main plots in July 1993 at the rates of 0, 2, 4, and 6 Mg ha-1. In June 2000, the main plots were divided in two subplots to study of the effect of surface re-liming at the rates of 0 and 3 Mg ha-1. The cropping sequence was soybean [Glycine max (L.) Merrill] (2001-2 and 2002-3), wheat (Triticum aestivum L.) (2003), soybean (2003-4), corn (Zea mays L.) (2004-5), and soybean (2005-6). Soil samples were collected at the following depths: 0-0.05, 0.05-0.10, and 0.10-0.20m, 10 years after surface liming and 3 years after surface re-liming. Soil Zn levels were extracted by four extractants: (i) 0.005molL-1 diethylenetriaminepentaacetic acid (DTPA) + 0.1molL-1 triethanolamine (TEA) + 0.01molL-1 calcium chloride (CaCl2) solution at pH7.3 (DTPA-TEA), (ii) 0.1molL-1 hydrochloric acid (HCl) solution, (iii) Mehlich 1 solution, and (iv) Mehlich 3 solution. Zinc concentrations in leaves and grains of soybean, wheat, and corn were also determined. Soil pH (0.01molL-1 CaCl2 suspension) varied from 4.4 to 6.1, at the 0- to 0.05-m depth, from 4.2 to 5.3 at the 0.05- to 0.10-m depth, and from 4.2 to 4.8 at the 0.10- to 0.20-m depth, after liming and re-liming. Zinc concentrations evaluated by DTPA-TEA, 0.1molL-1 HCl, Mehlich 1, and Mehlich 3 solutions were not changed as a result of lime rate application. Re-liming increased Zn concentrations extracted by 0.1molL-1 HCl at 0-0.05m deep and by DTPA-TEA at 0.05-0.10m deep. Surface-applied lime promoted a decrease in Zn concentrations of the crops, mainly in grains, because of increased soil pH at the surface layers. Regardless of the liming treatments, levels of Zn were sufficient to soybean, wheat, and corn nutrition under NT.

Podzolization as a deferralitization process: dynamics and chemistry of ground and surface waters in an Acrisol-Podzol sequence of the upper Amazon Basin

Hydrochemical processes involved in the development of hydromorphic Podzols are a major concern for the upper Amazon Basin because of the extent of the areas affected by such processes and the large amounts of organic carbon and associated metals exported to the rivers. The dynamics and chemical composition of ground and surface waters were studied along an Acrisol-Podzol sequence lying in an open depression of a plateau. Water levels were monitored along the sequence over a period of 2 years by means of piezometers. Water was sampled in zero-tension lysimeters for groundwater and for surface water in the drainage network of the depression. The pH and concentrations of organic carbon and major elements (Si, Fe and Al) were determined. The contrasted changes reported for concentrations of Si, organic carbon and metals (Fe, Al) mainly reflect the dynamics of the groundwater and the weathering conditions that prevail in the soils. Iron is released by the reductive dissolution of Fe oxides, mostly in the Bg horizons of the upslope Acrisols. It moves laterally under the control of hydraulic gradients and migrates through the iron-depleted Podzols where it is exported to the river network. Aluminium is released from the dissolution of Al-bearing minerals (gibbsite and kaolinite) at the margin of the podzolic area but is immobilized as organo-Al complexes in spodic horizons. In downslope positions, the quick recharge of the groundwater and large release of organic compounds lead to acidification and a loss of metals (mainly Al), previously stored in the Podzols.

A lipocalin sequence signature modulates cell survival

Lipocalins are beta-barrel proteins, which share three conserved motifs in their amino acid sequence. In this study, we identified by a peptide mapping approach, a seven-amino acid sequence related to one of these motifs (motif 2) that modulates cell survival. A synthetic peptide based on an insect lipocalin displayed cytoprotective activity in serum-deprived endothelial cells and leucocytes. This activity was dependent on nitric oxide synthase. This sequence was found within several lipocalins, including apolipoprotein D, retinol binding protein, lipocalin-type prostaglandin D synthase, and many unknown proteins, suggesting that it is a sequence signature and a lipocalin conserved property. (C) 2010 Federation of European Biochemical Societies. Published by Elsevier B. V. All rights reserved.