Overcoming function annotation errors in the Gram-positive pathogen Streptococcus suis by a proteomics-driven approach

Background: Annotation of protein-coding genes is a key step in sequencing projects. Protein functions are mainly assigned on the basis of the amino acid sequence alone by searching of homologous proteins. However, fully automated annotation processes often lead to wrong prediction of protein functions, and therefore time-intensive manual curation is often essential. Here we describe a fast and reliable way to correct function annotation in sequencing projects, focusing on surface proteomes. We use a proteomics approach, previously proven to be very powerful for identifying new vaccine candidates against Gram-positive pathogens. It consists of shaving the surface of intact cells with two proteases, the specific cleavage-site trypsin and the unspecific proteinase K, followed by LC/MS/MS analysis of the resulting peptides. The identified proteins are contrasted by computational analysis and their sequences are inspected to correct possible errors in function prediction. Results: When applied to the zoonotic pathogen Streptococcus suis, of which two strains have been recently sequenced and annotated, we identified a set of surface proteins without cytoplasmic contamination: all the proteins identified had exporting or retention signals towards the outside and/or the cell surface, and viability of protease-treated cells was not affected. The combination of both experimental evidences and computational methods allowed us to determine that two of these proteins are putative extracellular new adhesins that had been previously attributed a wrong cytoplasmic function. One of them is a putative component of the pilus of this bacterium. Conclusion: We illustrate the complementary nature of laboratory-based and computational methods to examine in concert the localization of a set of proteins in the cell, and demonstrate the utility of this proteomics-based strategy to experimentally correct function annotation errors in sequencing projects. This approach also contributes to provide strong experimental evidences that can be used to annotate those proteins for which a Gene Ontology (GO) term has not been assigned so far. Function annotation correction would then improve the identification of surfaceassociated proteins in bacterial pathogens, thus accelerating the discovery of new vaccines in infectious disease research

Conservation Agriculture and its contribution to the achievement of agri-environmental and economic challenges in Europe

Conservation Agriculture is an ecosystem approach to farming capable of providing solutions for numerous of the agri-environmental concerns in Europe. Certainly, most of the challenges addressed in the Common Agriculture Policy (CAP) could be tackled through Conservation Agriculture (CA). Not only the agri-environmental ones, but also those concerning farmer and rural communities’ prosperity. The optimisation of inputs and similar yields than conventional tillage, make Conservation Agriculture a profitable system compared to the tillage based agriculture. Whereas this sustainable agricultural system was conceived for protecting agrarian soils from its degradation, the numerous collateral benefits that emanate from soil conservation, i.e., climate change mitigation and adaptation, have raised Conservation Agriculture as one of the global emerging agrosciences, being adopted by an increasing number of farmers worldwide, including Europe

Efficacy of biosecurity measures in the control of microorganisms associated to endometritis in sows. Preliminary study

Biosecurity can be defined as all the applied measurements that take as a target to minimize the sanitary risks in a stock farm, and include measurements related to the facilities and the management. The efficacy of these measurements must be reflected in a decrease of the microorganism in different productive phases. A study was carried out to evaluate if the set of applied measurements influences the microbial uterine contamination after farrowing of healthy sows. Two swine farms were been completed about biosecurity measurements was completed and a microbiological study of uterine swabs of sows after the farrowing was carried out. A total of 60 animals were studied, and 27 (45%, 95% CI [33.3%, 56.7%]) resulted positive. Significant differences between production and selection and multiplication farms were detected (OR = 3.44, IC 95%, 1.135-11.047). The colonization frequency was 65% CI [51.3%, 78.6%] and 35% CI [21%, 49%] in production and selection farm, respectively (P = 0.02). A total of 66 isolates were obtained, represented mainly by Staphylococcus spp. (33.33%) and Aerococcus spp. (27.27%), although other species included in the genus Streptococcus (9.09%), Enterococcus (6.06%) and Pseudomonas (4.55%), as well as different fungi species were also isolated. The frequency of isolation of different microorganisms was similar in both farms, with the exception of the genus Enterococcus that was not isolated in the production farm (P = 0.01). The questionnaire showed some differences in biosecurity measures in the selection and multiplication farm when it is compared to the production farm, which together with the increased uterine microbial contamination observed in the latter leads us to propose a preliminary hypothesis about the possible risk factors associated with this process, highlighting the absence of measures to avoid the presence of vectors and the establishment of strict protocols for cleaning and disinfectio

Parameterisation and Optimisation of a Hand-Rake Sweeper: Application in Olive Picking

Olive picking is one of the most common social agricultural activities in many regions of Andalusia where the predominant crop is the traditional olive grove. The machinery used includes shakers, blowers and essential, low-cost hand-rake sweepers. The latter are generally used by the women of the squads to sweep the olives that fall from the trees. This article is focused on the design and optimisation of a hand-rake sweeper, in terms of durability and cost, for the picking of olives and other fruits, such as almonds, which are currently the main alternative to nonperennial crops in Andalusia. A parametric design of a hand-rake sweeper was created for this application using the design software CATIA, and its most vulnerable points were analysed in terms of effectiveness with varying design parameters, conducting usage simulations with ANSYS for a light material such as polypropylene. The maximum von Mises stress of the whole structure was 155.81 MPa. Using ANSYS, the dimension parameters of the hand-rake sweeper structure were optimised. The modified design was analysed again, showing a reduction of maximum tensions of 10.06%, as well as a decrease in its maximum elongations (0.0181 mm)

Risk factors associated with the antimicrobial resistance of staphylococci in canine pyoderma

This study reports the susceptibility to antimicrobial agents of staphylococci 4 (n=105) isolated from dogs, and the factors associated with this resistance. The study 5 animals were 23 healthy dogs (group A), 24 with first-time pyoderma (group B), and 27 6 with recurrent pyoderma that had undergone long-term antibiotic treatment (group C). 7 Staphylococci were more commonly isolated from the pyoderma-affected than the 8 healthy dogs (p<0.0001). 9 10 Some 78% of the isolates were resistant to at least one antimicrobial agent. 11 Resistance to amoxicillin-clavulanate,cephalosporins (OR 4.29, 95% CI [1.15, 16.3] 12 respectively), enrofloxacin (OR 9.47, 95% CI [1.53, 58.5]) and ciprofloxacin (OR 79.7 13 95% CI [3.26, 1947.4]) was more common among group C isolates. Some 32% of all 14 the isolates were multiresistant (MR) and 10.4% were methicillin-resistant (MRS). The 15 probability of isolating MRS staphylococci in group C increased by a factor of four 16 (95% CI [1.18, 17.9]) compared to A plus B. Multi-resistant (MR) isolates were 17 obtained more commonly from urban than rural dogs (OR 3.79, 95% CI [1.09, 13.17]). 18 All the MRS staphylococci encountered were obtained from urban dogs and more 19 commonly from male dogs (p= 0.07). 20 21 This study shows that dogs bred in urban habitat, with a history of antibiotic 22 therapy in the past year represents significant risk of being carriers of isolates resistant 23 to methicillin (MRS) and other antimicrobials. These factors should be considered 24 before applying an antimicrobial treatment in veterinary clinics

Search of Potential Vaccine Candidates against Trueperella pyogenes Infections through Proteomic and Bioinformatic Analysis

Trueperella pyogenes is an opportunistic pathogen, responsible for important infections in pigs and significant economic losses in swine production. To date, there are no available commercial vaccines to control diseases caused by this bacterium. In this work, we performed a comparative proteomic analysis of 15 T. pyogenes clinical isolates, by “shaving” live cells, followed by LC-MS/MS, aiming at the identification of the whole set of surface proteins (i.e., the “pan-surfome”) as a source of antigens to be tested in further studies as putative vaccine candidates, or used in diagnostic tools. A total of 140 surface proteins were detected, comprising 25 cell wall proteins, 10 secreted proteins, 23 lipoproteins and 82 membrane proteins. After describing the “pan-surfome”, the identified proteins were ranked in three different groups based on the following criteria: to be (i) surface-exposed, (ii) highly conserved and (iii) widely distributed among different isolates. Two cell wall proteins, three lipoproteins, four secreted and seven membrane proteins were identified in more than 70% of the studied strains, were highly expressed and highly conserved. These proteins are potential candidates, alone or in combination, to obtain effective vaccines against T. pyogenes or to be used in the diagnosis of this pathogen

Identification of new immunoprotective surface protein vaccine candidates against streptococcus suis infection by proteomics

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A Novel Missense Mutation, I890T, in the Pore Region of Cardiac Sodium Channel Causes Brugada Syndrome

Brugada syndrome (BrS) is a life-threatening, inherited arrhythmogenic syndrome associated with autosomal dominant mutations in SCN5A, the gene encoding the cardiac Na+ channel alpha subunit (Nav1.5). The aim of this work was to characterize the functional alterations caused by a novel SCN5A mutation, I890T, and thus establish whether this mutation is associated with BrS. The mutation was identified by direct sequencing of SCN5A from the proband's DNA. Wild-type (WT) or I890T Nav1.5 channels were heterologously expressed in human embryonic kidney cells. Sodium currents were studied using standard whole cell patch-clamp protocols and immunodetection experiments were performed using an antibody against human Nav1.5 channel. A marked decrease in current density was observed in cells expressing the I890T channel (from -52.0±6.5 pA/pF, n = 15 to -35.9±3.4 pA/pF, n = 22, at -20 mV, WT and I890T, respectively). Moreover, a positive shift of the activation curve was identified (V1/2 = -32.0±0.3 mV, n = 18, and -27.3±0.3 mV, n = 22, WT and I890T, respectively). No changes between WT and I890T currents were observed in steady-state inactivation, time course of inactivation, slow inactivation or recovery from inactivation parameters. Cell surface protein biotinylation analyses confirmed that Nav1.5 channel membrane expression levels were similar in WT and I890T cells. In summary, our data reveal that the I890T mutation, located within the pore of Nav1.5, causes an evident loss-of-function of the channel. Thus, the BrS phenotype observed in the proband is most likely due to this mutation. © 2013 Tarradas et al

Making Climate Change Mitigation and Adaptability Real in Africa with Conservation Agriculture

In this report, the authors have gathered essential information on how the agricultural sector can respond to climate change through Conservation Agriculture (CA). This document aims to serve as a basis for decision-making based on science and agricultural experimentation in Africa

Challenge of Pigs with Classical Swine Fever Viruses after C-Strain Vaccination Reveals Remarkably Rapid Protection and Insights into Early Immunity

Pre-emptive culling is becoming increasingly questioned as a means of controlling animal diseases, including classical swine fever (CSF). This has prompted discussions on the use of emergency vaccination to control future CSF outbreaks in domestic pigs. Despite a long history of safe use in endemic areas, there is a paucity of data on aspects important to emergency strategies, such as how rapidly CSFV vaccines would protect against transmission, and if this protection is equivalent for all viral genotypes, including highly divergent genotype 3 strains. To evaluate these questions, pigs were vaccinated with the Riemser® C-strain vaccine at 1, 3 and 5 days prior to challenge with genotype 2.1 and 3.3 challenge strains. The vaccine provided equivalent protection against clinical disease caused by for the two challenge strains and, as expected, protection was complete at 5 days post-vaccination. Substantial protection was achieved after 3 days, which was sufficient to prevent transmission of the 3.3 strain to animals in direct contact. Even by one day post-vaccination approximately half the animals were partially protected, and were able to control the infection, indicating that a reduction of the infectious potential is achieved very rapidly after vaccination. There was a close temporal correlation between T cell IFN-γ responses and protection. Interestingly, compared to responses of animals challenged 5 days after vaccination, challenge of animals 3 or 1 days post-vaccination resulted in impaired vaccine-induced T cell responses. This, together with the failure to detect a T cell IFN-γ response in unprotected and unvaccinated animals, indicates that virulent CSFV can inhibit the potent antiviral host defences primed by C-strain in the early period post vaccination