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Vibrio alginolyticus is a Gram-negative bacterium commonly associated with mackerel poisoning. A bacteriophage that specifically targets and lyses this bacterium could be employed as a biocontrol agent for treating the bacterial infection or improving the shelf-life of mackerel products. However, only a few well-characterized V. alginolyticus phages have been reported in the literature. In this study, a novel lytic phage, named ΦImVa-1, specifically infecting V. alginolyticus strain ATCC 17749, was isolated from Indian mackerel. The phage has a short latent period of 15 min and a burst size of approximately 66 particles per infected bacterium. ΦImVa-1 remained stable for 2 h at a wide temperature (27-75 °C) and within a pH range of 5 to 10. Transmission electron microscopy revealed that ΦImVa-1 has an icosahedral head of approximately 60 nm in diameter with a short tail, resembling those in the Schitoviridae family. High throughput sequencing and bioinformatics analysis elucidated that ΦImVa-1 has a linear dsDNA genome of 77,479 base pairs (bp), with a G + C content of ~ 38.72% and 110 predicted gene coding regions (106 open reading frames and four tRNAs). The genome contains an extremely large virion-associated RNA polymerase gene and two smaller non-virion-associated RNA polymerase genes, which are hallmarks of schitoviruses. No antibiotic genes were found in the ΦImVa-1 genome. This is the first paper describing the biological properties, morphology, and the complete genome of a V. alginolyticus-infecting schitovirus. When raw mackerel fish flesh slices were treated with ΦImVa-1, the pathogen loads reduced significantly, demonstrating the potential of the phage as a biocontrol agent for V. alginolyticus strain ATCC 17749 in the food. KEY POINTS: • A novel schitovirus infecting Vibrio alginolyticus ATCC 17749 was isolated from Indian mackerel. • The complete genome of the phage was determined, analyzed, and compared with other phages. • The phage is heat stable making it a potential biocontrol agent in extreme environments.

The etiological agent of melioidosis, Burkholderia pseudomallei, utilises a type III secretion system cluster 3 (T3SS3) to deliver proteins termed type III effectors (T3SEs) into the host cytoplasm in order to establish an intracellular lifecycle in phagocytic and non-phagocytic cells, thus playing an important role in pathogenesis. To gain insight into possible functional roles for BapA, a putative T3SE with unknown function, in the intracellular lifecycle of B.pseudomallei, bapA gene knockout mutant was constructed. The effect of the knockout on virulence to the otherwise isogenic parental strain, K96243, was studied by cellular infection assays and Caenorhabditis elegans killing assay. The attachment and subsequent entry into A549 cells was significantly (P < 0.05) attenuated in the Delta bapA compared to K96243. However, intracellular replication was not affected. Furthermore, the cell-to-cell spreading capacity of Delta bapA was impaired although the mutant exhibited no evident defect in its actin tail formation. Additionally, phagocytosis and intracellular replication rates of Delta bapA in U937 macrophage cells were significantly reduced relative to K96243 without phagosomal escape being affected. Based on these observations, we conclude that the BapA T3SE could play an important role in B.pseudomallei intracellular lifecycle, especially, in the early stages of attachment and entry into the host cell.

In the onshore drilling operation the main objective is always finding ways to optimize cost and improve the efficiency of drilling operations. Among the various available option, one possibility was to drill 17.5" deviated section in one run through the interbedded formation, which cause high vibrations and risk of twist-off. This section previously was drilled with minimum 2-3 bit runs for a heavy casing design. This would definitely reduce the well duration and cost. The plan involved to drill 17.5" deviated section using rotary steerable system using hybrid bit technology. Recent advances in drilling bit design has proved to be very effective in drilling surface hole sections but are limited to drill vertical holes and require multiple runs to complete a section. Special design and cutting structure is required when it comes to drill deviated hole. One supplier has combined the traditional design and come up with hybrid bit structure to achieve this goal of drilling surface deviated hole in one run. This special hybrid bit, drilled successfully 17.5" deviated section in one run with enhanced ROP by 40% compared to previous wells. This saved additional trips to change bit and avoided any stuck pipe and twist off. This kind of strategy has helped to maximize average ROP of 64 ft/hr for the entire section. The main element in optimizing the performance of is the systematic approach towards the bit selection, hydraulics and mud parameters. Outcome of this optimization resulted in case history data which shows that this kind of hybrid bit technology can be used to drill deviated wellbore with better penetration rates, lesser washouts and longer on-bottom time. This technical paper describes the results of first well drilled by a service provider using hybrid bit technology with rotary steerable system in one run. This has resulted in increasing the rate of penetration for the 17.5" deviated top hole section. Applying this kind of hybrid bit technology has not only enhanced the ROP but also helped to save rig days and cost.

MoSwA: Protein Sequence Diversity Motif Switch Analyser for Viruses

Avian influenza virus A (H7N9), after circulating in avian hosts for decades, was identified as a human pathogen in 2013. Herein, amino acid substitutions possibly essential for human adaptation were identified by comparing the 4706 aligned overlapping nonamer position sequences (1-9, 2-10, etc.) of the reported 2014 and 2017 avian and human H7N9 datasets. The initial set of virus sequences (as of year 2014) exhibited a total of 109 avian-to-human (A2H) signature amino acid substitutions. Each represented the most prevalent substitution at a given avian virus nonamer position that was selectively adapted as the corresponding index (most prevalent sequence) of the human viruses. The majority of these avian substitutions were long-standing in the evolution of H7N9, and only 17 were first detected in 2013 as possibly essential for the initial human adaptation. Strikingly, continued evolution of the avian H7N9 virus has resulted in avian and human protein sequences that are almost identical. This rapid and continued adaptation of the avian H7N9 virus to the human host, with near identity of the avian and human viruses, is associated with increased human infection and a predicted greater risk of human-to-human transmission. Bezmiâlem Vakıf Üniversitesi Avian Influenza H7N9 Virus Adaptation to Human Hosts

Background Biology has entered the era of big data with the advent of high-throughput omics technologies. Biological databases provide public access to petabytes of data and information facilitating knowledge discovery. Over the years, sequence data of pathogens has seen a large increase in the number of records, given the relatively small genome size and their important role as infectious and symbiotic agents. Humans are host to numerous pathogenic diseases, such as that by viruses, many of which are responsible for high mortality and morbidity. The interaction between pathogens and humans over the evolutionary history has resulted in sharing of sequences, with important biological and evolutionary implications. Results This study describes a large-scale, systematic bioinformatics approach for identification and characterization of shared sequences between the host and pathogen. An application of the approach is demonstrated through identification and characterization of the Flaviviridae-human share-ome. A total of 2430 nonamers represented the Flaviviridae-human share-ome with 100% identity. Although the share-ome represented a small fraction of the repertoire of Flaviviridae (~ 0.12%) and human (~ 0.013%) non-redundant nonamers, the 2430 shared nonamers mapped to 16,946 Flaviviridae and 7506 human non-redundant protein sequences. The shared nonamer sequences mapped to 125 species of Flaviviridae, including several with unclassified genus. The majority (~ 68%) of the shared sequences mapped to Hepacivirus C species; West Nile, dengue and Zika viruses of the Flavivirus genus accounted for ~ 11%, ~ 7%, and ~ 3%, respectively, of the Flaviviridae protein sequences (16,946) mapped by the share-ome. Further characterization of the share-ome provided important structural-functional insights to Flaviviridae-human interactions. Conclusion Mapping of the host-pathogen share-ome has important implications for the design of vaccines and drugs, diagnostics, disease surveillance and the discovery of unknown, potential host-pathogen interactions. The generic workflow presented herein is potentially applicable to a variety of pathogens, such as of viral, bacterial or parasitic origin.

In the current era of Big Data production and Artificial Intelligence development, one of the fastest growing scientific and professional areas that is generating petabytes of data is Life Sciences. In this context, many scientific and educational institutions recognize that it is no longer possible to carry out adequate studies and research in this area without well-trained computational biologists and bioinformaticians. GOBLET is an international organization, established in 2012, with the mission to cultivate a global community of bioinformatics trainers who support learning, education and training. GOBLET's mission was defined under the vision that there is a clear worldwide need to harmonize bioinformatics training activities and to unite, inspire and equip bioinformatics trainers. This can only be achieved by developing active linkages and collaborations with many national and international institutions that work in the field of Life Sciences. Relevant activities conducted last year by GOBLET members, together with other international experts, include the initiation of a series of assets, standards and guidelines, to define competencies, promote best practices and provide high-quality resources for learning, education and training in bioinformatics and computational biology, worldwide. GOBLET's new website offers a comprehensive portal, providing many materials and resources for the international bioinformatics community.

The sequence diversity of dengue virus (DENV) is one of the challenges in developing an effective vaccine against the virus. Highly conserved, serotype-specific (HCSS), immune-relevant DENV sequences are attractive candidates for vaccine design, and represent an alternative to the approach of selecting pan-DENV conserved sequences. The former aims to limit the number of possible cross-reactive epitope variants in the population, while the latter aims to limit the cross-reactivity between the serotypes to favour a serotype-specific response. Herein, we performed a large-scale systematic study to map and characterise HCSS sequences in the DENV proteome.All reported DENV protein sequence data for each serotype was retrieved from the NCBI Entrez Protein (nr) Database (txid: 12637). The downloaded sequences were then separated according to the individual serotype proteins by use of BLASTp search, and subsequently removed for duplicates and co-aligned across the serotypes. Shannon's entropy and mutual information (MI) analyses, by use of AVANA, were performed to measure the diversity within and between the serotype proteins to identify HCSS nonamers. The sequences were evaluated for the presence of promiscuous T-cell epitopes by use of NetCTLpan 1.1 and NetMHCIIpan 3.2 server for human leukocyte antigen (HLA) class I and class II supertypes, respectively. The predicted epitopes were matched to reported epitopes in the Immune Epitope Database.A total of 2321 nonamers met the HCSS selection criteria of entropy 0.25 and MI 0.8. Concatenating these resulted in a total of 337 HCSS sequences. DENV4 had the most number of HCSS nonamers; NS5, NS3 and E proteins had among the highest, with none in the C and only one in prM. The HCSS sequences were immune-relevant; 87 HCSS sequences were both reported T-cell epitopes/ligands in human and predicted epitopes, supporting the accuracy of the predictions. A number of the HCSS clustered as immunological hotspots and exhibited putative promiscuity beyond a single HLA supertype. The HCSS sequences represented, on average, ~ 40% of the proteome length for each serotype; more than double of pan-DENV sequences (conserved across the four serotypes), and thus offer a larger choice of sequences for vaccine target selection. HCSS sequences of a given serotype showed significant amino acid difference to all the variants of the other serotypes, supporting the notion of serotype-specificity.This work provides a catalogue of HCSS sequences in the DENV proteome, as candidates for vaccine target selection. The methodology described herein provides a framework for similar application to other pathogens.

Plasma leakage is the main pathophysiological feature in severe dengue, resulting from altered vascular barrier function associated with an inappropriate immune response triggered upon infection. The present study investigated functional changes using an electric cell-substrate impedance sensing system in four (brain, dermal, pulmonary and retinal) human microvascular endothelial cell (MEC) lines infected with purified dengue virus, followed by assessment of cytokine profiles and the expression of inter-endothelial junctional proteins. Modelling of changes in electrical impedance suggests that vascular leakage in dengue-infected MECs is mostly due to the modulation of cell-to-cell interactions, while this loss of vascular barrier function observed in the infected MECs varied between cell lines and DENV serotypes. High levels of inflammatory cytokines (IL-6 and TNF-alpha), chemokines (CXCL1, CXCL5, CXCL11, CX3CL1, CCL2 and CCL20) and adhesion molecules (VCAM-1) were differentially produced in the four infected MECs. Further, the tight junctional protein, ZO-1, was down-regulated in both the DENV-1-infected brain and pulmonary MECs, while claudin-1, PECAM-1 and VE-cadherin were differentially expressed in these two MECs after infection. Non-purified virus stock was also studied to investigate the impact of virus stock purity on dengue-specific immune responses, and the results suggest that virus stock propagated through cell culture may include factors that mask or alter the DENV-specific immune responses of the MECs. The findings of the present study show that high DENV load differentially modulates human microvascular endothelial barrier function and disrupts the function of inter-endothelial junctional proteins during early infection with organ-specific cytokine production.

Gas Injector Well is part of the project to improve the pressure communication between peripheral gas injectors and nearby producers in the Northern Part of Unit G WAG Patterns. This well was injecting gas only to upper part of Unit G reservoir based on the PLT and TGT results. Based on the study using tracer chemical injection, this upper Unit G reservoir was found to be in connecting with another nearby different reservoir. The initial plan was to re-horizontalize the well and place injector in the proper reservoir, later this was changed and revised to use this innovation technology by installing blank liner with isolating packer to isolate the upper part of Unit G reservoir and let the lower part of open-hole remained open and connected to the required layer of reservoir. After extensive discussion with vendors and performing simulation runs, finally agreed to run this kind of swell packer. Vendor custom designed this kind of innovative packer at their USA facility and transported to ADNOC Onshore yard just before the execution phase. First run the swell packer on blank pipe and placed it at desired depth in the open hole, later run upper completions and sting into the top packer of lower completion. This way, we were able to inject gas into the lower part of reservoir Unit-G only, whereas the upper part was remained isolated completely. Using this technique saved company additional 2 million and extra time for re-horizontalization.

Background Ebolavirus (EBOV) is responsible for one of the most fatal diseases encountered by mankind. Cellular T-cell responses have been implicated to be important in providing protection against the virus. Antigenic variation can result in viral escape from immune recognition. Mapping targets of immune responses among the sequence of viral proteins is, thus, an important first step towards understanding the immune responses to viral variants and can aid in the identification of vaccine targets. Herein, we performed a large-scale, proteome-wide mapping and diversity analyses of putative HLA supertype-restricted T-cell epitopes of Zaire ebolavirus (ZEBOV), the most pathogenic species among the EBOV family. Methods All publicly available ZEBOV sequences (14,098) for each of the nine viral proteins were retrieved, removed of irrelevant and duplicate sequences, and aligned. The overall proteome diversity of the non-redundant sequences was studied by use of Shannon’s entropy. The sequences were predicted, by use of the NetCTLpan server, for HLA-A2, -A3, and -B7 supertype-restricted epitopes, which are relevant to African and other ethnicities and provide for large (~86%) population coverage. The predicted epitopes were mapped to the alignment of each protein for analyses of antigenic sequence diversity and relevance to structure and function. The putative epitopes were validated by comparison with experimentally confirmed epitopes. Results & discussion ZEBOV proteome was generally conserved, with an average entropy of 0.16. The 185 HLA supertype-restricted T-cell epitopes predicted (82 (A2), 37 (A3) and 66 (B7)) mapped to 125 alignment positions and covered ~24% of the proteome length. Many of the epitopes showed a propensity to co-localize at select positions of the alignment. Thirty (30) of the mapped positions were completely conserved and may be attractive for vaccine design. The remaining (95) positions had one or more epitopes, with or without non-epitope variants. A significant number (24) of the putative epitopes matched reported experimentally validated HLA ligands/T-cell epitopes of A2, A3 and/or B7 supertype representative allele restrictions. The epitopes generally corresponded to functional motifs/domains and there was no correlation to localization on the protein 3D structure. These data and the epitope map provide important insights into the interaction between EBOV and the host immune system. Electronic supplementary material The online version of this article (10.1186/s12864-017-4328-8) contains supplementary material, which is available to authorized users.

SUMOylation is a form of post-translational modification involving covalent attachment of SUMO (Small Ubiquitin-like Modifier) polypeptides to specific lysine residues in the target protein. In human cells, there are four SUMO proteins, SUMO1-4, with SUMO2 and SUMO3 forming a closely related subfamily. SUMO2/3, in contrast to SUMO1, are predominantly involved in the cellular response to certain stresses, including heat shock. Substantial evidence from studies in yeast has shown that SUMOylation plays an important role in the regulation of DNA replication and repair. Here, we report a proteomic analysis of proteins modified by SUMO2 in response to DNA replication stress in S phase in human cells. We have identified a panel of 22 SUMO2 targets with increased SUMOylation during DNA replication stress, many of which play key functions within the DNA replication machinery and/or in the cellular response to DNA damage. Interestingly, POLD3 was found modified most significantly in response to a low dose aphidicolin treatment protocol that promotes common fragile site (CFS) breakage. POLD3 is the human ortholog of POL32 in budding yeast, and has been shown to act during break-induced recombinational repair. We have also shown that deficiency of POLD3 leads to an increase in RPA-bound ssDNA when cells are under replication stress, suggesting that POLD3 plays a role in the cellular response to DNA replication stress. Considering that DNA replication stress is a source of genome instability, and that excessive replication stress is a hallmark of pre-neoplastic and tumor cells, our characterization of SUMO2 targets during a perturbed S-phase should provide a valuable resource for future functional studies in the fields of DNA metabolism and cancer biology.

Background Viral vaccine target discovery requires understanding the diversity of both the virus and the human immune system. The readily available and rapidly growing pool of viral sequence data in the public domain enable the identification and characterization of immune targets relevant to adaptive immunity. A systematic bioinformatics approach is necessary to facilitate the analysis of such large datasets for selection of potential candidate vaccine targets. Results This work describes a computational methodology to achieve this analysis, with data of dengue, West Nile, hepatitis A, HIV-1, and influenza A viruses as examples. Our methodology has been implemented as an analytical pipeline that brings significant advancement to the field of reverse vaccinology, enabling systematic screening of known sequence data in nature for identification of vaccine targets. This includes key steps (i) comprehensive and extensive collection of sequence data of viral proteomes (the virome), (ii) data cleaning, (iii) large-scale sequence alignments, (iv) peptide entropy analysis, (v) intra- and inter-species variation analysis of conserved sequences, including human homology analysis, and (vi) functional and immunological relevance analysis. Conclusion These steps are combined into the pipeline ensuring that a more refined process, as compared to a simple evolutionary conservation analysis, will facilitate a better selection of vaccine targets and their prioritization for subsequent experimental validation. Electronic supplementary material The online version of this article (10.1186/s12920-017-0301-2) contains supplementary material, which is available to authorized users.

Efficient High Power Microwave radiating systems are required to extract high power microwaves from HPM devices. Efficiency of an HPM antenna can be improved by varying its physical shape and dielectric used in it. The most critical location in an HPM antenna is its interface from where microwave leaves and enters into the air. In this paper, the performance of an HPM antenna has been investigated as a function of its profile. It has been demonstrated that by adopting Gaussian beam distribution at the aperture of an horn antenna with an appropriate beam waist, the E-field intensity at the output window can significantly be reduced resulting into 29 % improvement in its gain. Thus, in general, improved characteristics of an HPM antenna can be achieved by adopting an appropriate horn profile, beam waist and dielectric.

In this work, a high power microwave (HPM) antenna is designed and its breakdown limits are investigated. It has been shown that the designed antenna can handle Giga Watt (GW) power without attaining breakdown. The flash-over which usually occurs at the interface of HPM antennas, where microwave leaves and enters into the air, was controlled by adjusting the field intensity at the output window of the antenna. It has been shown that by adopting Gaussian beam distribution at the aperture of horn with appropriate beam waist, the E-field intensity at the output window can be reduced significantly. It has also been demonstrated that an HPM antenna with 0.4 m aperture radius can sustain >30 GW power for a pulse duration of 1ns and the same is reduced to 1μs.

West Nile virus (WNV) has emerged globally as an increasingly important pathogen for humans and domestic animals. Studies of the evolutionary diversity of the virus over its known history will help to elucidate conserved sites, and characterize their correspondence to other pathogens and their relevance to the immune system. We describe a large-scale analysis of the entire WNV proteome, aimed at identifying and characterizing evolutionarily conserved amino acid sequences. This study, which used 2,746 WNV protein sequences collected from the NCBI GenPept database, focused on analysis of peptides of length 9 amino acids or more, which are immunologically relevant as potential T-cell epitopes. Entropy-based analysis of the diversity of WNV sequences, revealed the presence of numerous evolutionarily stable nonamer positions across the proteome (entropy value of < or = 1). The representation (frequency) of nonamers variant to the predominant peptide at these stable positions was, generally, low (< or = 10% of the WNV sequences analyzed). Eighty-eight fragments of length 9-29 amino acids, representing approximately 34% of the WNV polyprotein length, were identified to be identical and evolutionarily stable in all analyzed WNV sequences. Of the 88 completely conserved sequences, 67 are also present in other flaviviruses, and several have been associated with the functional and structural properties of viral proteins. Immunoinformatic analysis revealed that the majority (78/88) of conserved sequences are potentially immunogenic, while 44 contained experimentally confirmed human T-cell epitopes. This study identified a comprehensive catalogue of completely conserved WNV sequences, many of which are shared by other flaviviruses, and majority are potential epitopes. The complete conservation of these immunologically relevant sequences through the entire recorded WNV history suggests they will be valuable as components of peptide-specific vaccines or other therapeutic applications, for sequence-specific diagnosis of a wide-range of Flavivirus infections, and for studies of homologous sequences among other flaviviruses.

Phylogenetic relatedness and cocirculation of several major human pathogen flaviviruses are recognized as a possible cause of deleterious immune responses to mixed infection or immunization and call for a greater understanding of the inter- Flavivirus protein homologies. This study focused on the identification of human leukocyte antigen (HLA)-restricted West Nile virus (WNV) T-cell ligands and characterization of their distribution in reported sequence data of WNV and other flaviviruses. H-2-deficient mice transgenic for either A2, A24, B7, DR2, DR3, or DR4 HLA alleles were immunized with overlapping peptides of the WNV proteome, and peptide-specific T-cell activation was measured by gamma interferon (IFN-γ) enzyme-linked immunosorbent spot (ELISpot) assays. Approximately 30% (137) of the WNV proteome peptides were identified as HLA-restricted T-cell ligands. The majority of these ligands were conserved in ∼≥88% of analyzed WNV sequences. Notably, only 51 were WNV specific, and the remaining 86, chiefly of E, NS3, and NS5, shared an identity of nine or more consecutive amino acids with sequences of 64 other flaviviruses, including several major human pathogens. Many of the shared ligands had an incidence of >50% in the analyzed sequences of one or more of six major flaviviruses. The multitude of WNV sequences shared with other flaviviruses as interspecies variants highlights the possible hazard of defective T-cell activation by altered peptide ligands in the event of dual exposure to WNV and other flaviviruses, by either infection or immunization. The data suggest the possible preferred use of sequences that are pathogen specific with minimum interspecies sequence homology for the design of Flavivirus vaccines.

African trypanosomes are tsetse fly transmitted protozoan parasites responsible for human African trypanosomiasis, a disease characterized by a plethora of neurological symptoms and death. How the parasites under microvascular shear stress (SS) flow conditions in the brain cross the blood-brain barrier (BBB) is not known. In vitro studies using static models comprised of human brain microvascular endothelial cells (BMEC) show that BBB activation and crossing by trypanosomes requires the orchestration of parasite cysteine proteases and host calcium-mediated cell signaling. Here, we examine BMEC barrier function and the activation of extracellular signal-regulated kinase (ERK)1/2 and ERK5, mitogen-activated protein kinase family regulators of microvascular permeability, under static and laminar SS flow and in the context of trypanosome infection. Confluent human BMEC were cultured in electric cell-substrate impedance sensing (ECIS) and parallel-plate glass slide chambers. The human BMEC were exposed to 2 or 14 dyn/cm(2) SS in the presence or absence of trypanosomes. Real-time changes in transendothelial electrical resistance (TEER) were monitored and phosphorylation of ERK1/2 and ERK5 analyzed by immunoblot assay. After reaching confluence under static conditions human BMEC TEER was found to rapidly increase when exposed to 2 dyn/cm(2) SS, a condition that mimics SS in brain postcapillary venules. Addition of African trypanosomes caused a rapid drop in human BMEC TEER. Increasing SS to 14 dyn/cm(2), a condition mimicking SS in brain capillaries, led to a transient increase in TEER in both control and infected human BMEC. However, no differences in ERK1/2 and ERK5 activation were found under any condition tested. African trypanosomiasis alters BBB permeability under low shear conditions through an ERK1/2 and ERK5 independent pathway.

Epitope-based vaccines provide a new strategy for prophylactic and therapeutic application of pathogen-specific immunity. A critical requirement of this strategy is the identification and selection of T-cell epitopes that act as vaccine targets. This study describes current methodologies for the selection process, with dengue virus as a model system. A combination of publicly available bioinformatics algorithms and computational tools are used to screen and select antigen sequences as potential T-cell epitopes of supertype human leukocyte antigen (HLA) alleles. The selected sequences are tested for biological function by their activation of T-cells of HLA transgenic mice and of pathogen infected subjects. This approach provides an experimental basis for the design of pathogen specific, T-cell epitope-based vaccines that are targeted to majority of the genetic variants of the pathogen, and are effective for a broad range of differences in human leukocyte antigens among the global human population.