331 research outputs found
The importance of the glycosylation of antimicrobial peptides: natural and synthetic approaches.
Glycosylation is one of the most prevalent post-translational modifications of a protein, with a defining impact on its structure and function. Many of the proteins involved in the innate or adaptive immune response, including cytokines, chemokines, and antimicrobial peptides (AMPs), are glycosylated, contributing to their myriad activities. The current availability of synthetic coupling and glycoengineering technology makes it possible to customise the most beneficial glycan modifications for improved AMP stability, microbicidal potency, pathogen specificity, tissue or cell targeting, and immunomodulation
Intracellular replication of the well-armed pathogen Burkholderia pseudomallei.
The Burkholderia genus contains a group of soil-dwelling Gram-negative organisms that are prevalent in warm and humid climates. Two species in particular are able to cause disease in animals, B. mallei primarily infects Equus spp. and B. pseudomallei (BPS), that is able to cause potentially life-threatening disease in humans. BPS is naturally resistant to many antibiotics and there is no vaccine available. Although not a specialised human pathogen, BPS possesses a large genome and many virulence traits that allow it to adapt and survive very successfully in the human host. Key to this survival is the ability of BPS to replicate intracellularly. In this review we highlight recent advances in our understanding of the intracellular survival of BPS, including how it overcomes host immune defenses and other challenges to establish its niche and then spread the infection. Knowledge of these mechanisms increases our capacity for therapeutic interventions against a well-armed foe
Virulence of the emerging pathogen, Burkholderia pseudomallei, depends upon the O-linked oligosaccharyltransferase, PglL.
Aim: We sought to characterize the contribution of the O-OTase, PglL, to virulence in two Burkholderia spp. by comparing isogenic mutants in Burkholderia pseudomallei with the related species, Burkholderia thailandensis. Materials & methods: We utilized an array of in vitro assays in addition to Galleria mellonella and murine in vivo models to assess virulence of the mutant and wild-type strains in each Burkholderia species. Results: We found that pglL contributes to biofilm and twitching motility in both species. PglL uniquely affected morphology; cell invasion; intracellular motility; plaque formation and intergenus competition in B. pseudomallei. This mutant was attenuated in the murine model, and extended survival in a vaccine-challenge experiment. Conclusion: Our data support a broad role for pglL in bacterial fitness and virulence, particularly in B. pseudomallei
High-throughput analysis of Yersinia pseudotuberculosis gene essentiality in optimised in vitro conditions, and implications for the speciation of Yersinia pestis.
BACKGROUND: Yersinia pseudotuberculosis is a zoonotic pathogen, causing mild gastrointestinal infection in humans. From this comparatively benign pathogenic species emerged the highly virulent plague bacillus, Yersinia pestis, which has experienced significant genetic divergence in a relatively short time span. Much of our knowledge of Yersinia spp. evolution stems from genomic comparison and gene expression studies. Here we apply transposon-directed insertion site sequencing (TraDIS) to describe the essential gene set of Y. pseudotuberculosis IP32953 in optimised in vitro growth conditions, and contrast these with the published essential genes of Y. pestis. RESULTS: The essential genes of an organism are the core genetic elements required for basic survival processes in a given growth condition, and are therefore attractive targets for antimicrobials. One such gene we identified is yptb3665, which encodes a peptide deformylase, and here we report for the first time, the sensitivity of Y. pseudotuberculosis to actinonin, a deformylase inhibitor. Comparison of the essential genes of Y. pseudotuberculosis with those of Y. pestis revealed the genes whose importance are shared by both species, as well as genes that were differentially required for growth. In particular, we find that the two species uniquely rely upon different iron acquisition and respiratory metabolic pathways under similar in vitro conditions. CONCLUSIONS: The discovery of uniquely essential genes between the closely related Yersinia spp. represent some of the fundamental, species-defining points of divergence that arose during the evolution of Y. pestis from its ancestor. Furthermore, the shared essential genes represent ideal candidates for the development of novel antimicrobials against both species
Amperometric and spectrophotometric determination of carbaryl in natural waters and commercial formulations
The work presented describes the development
and evaluation of two flow-injection analysis (FIA) systems
for the automated determination of carbaryl in spiked
natural waters and commercial formulations. Samples are
injected directly into the system where they are subjected
to alkaline hydrolysis thus forming 1-naphthol. This product
is readily oxidised at a glassy carbon electrode. The
electrochemical behaviour of 1-naphthol allows the development
of an FIA system with an amperometric detector
in which 1-naphthol determination, and thus measurement
of carbaryl concentration, can be performed. Linear response
over the range 1.0×10–7 to 1.0×10–5 mol L–1, with a
sampling rate of 80 samples h–1, was recorded. The detection
limit was 1.0×10–8 mol L–1. Another FIA manifold was
constructed but this used a colorimetric detector. The methodology
was based on the coupling of 1-naphthol with phenylhydrazine
hydrochloride to produce a red complex which
has maximum absorbance at 495 nm. The response was
linear from 1.0×10–5 to 1.5×10–3 mol L–1 with a detection
limit of 1.0×10–6 mol L–1. Sample-throughput was about
60 samples h–1. Validation of the results provided by the
two FIA methodologies was performed by comparing
them with results from a standard HPLC–UV technique.
The relative deviation was <5%. Recovery trials were also
carried out and the values obtained ranged from 97.0 to
102.0% for both methods. The repeatability (RSD, %) of
12 consecutive injections of one sample was 0.8% and
1.6% for the amperometric and colorimetric systems, respectively
A Method to Improve the Early Stages of the Robotic Process Automation Lifecycle
The robotic automation of processes is of much interest to
organizations. A common use case is to automate the repetitive manual
tasks (or processes) that are currently done by back-office staff
through some information system (IS). The lifecycle of any Robotic Process
Automation (RPA) project starts with the analysis of the process
to automate. This is a very time-consuming phase, which in practical
settings often relies on the study of process documentation. Such documentation
is typically incomplete or inaccurate, e.g., some documented
cases never occur, occurring cases are not documented, or documented
cases differ from reality. To deploy robots in a production environment
that are designed on such a shaky basis entails a high risk. This paper
describes and evaluates a new proposal for the early stages of an RPA
project: the analysis of a process and its subsequent design. The idea is to
leverage the knowledge of back-office staff, which starts by monitoring
them in a non-invasive manner. This is done through a screen-mousekey-
logger, i.e., a sequence of images, mouse actions, and key actions
are stored along with their timestamps. The log which is obtained in
this way is transformed into a UI log through image-analysis techniques
(e.g., fingerprinting or OCR) and then transformed into a process model
by the use of process discovery algorithms. We evaluated this method for
two real-life, industrial cases. The evaluation shows clear and substantial
benefits in terms of accuracy and speed. This paper presents the method,
along with a number of limitations that need to be addressed such that
it can be applied in wider contexts.Ministerio de EconomÃa y Competitividad TIN2016-76956-C3-2-
Re-sensitization of Mycobacterium smegmatis to Rifampicin Using CRISPR Interference Demonstrates Its Utility for the Study of Non-essential Drug Resistance Traits
© 2021 Faulkner, Cox, Goh, van Bohemen, Gibson, Liebster, Wren, Willcocks and Kendall. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). https://creativecommons.org/licenses/by/4.0/A greater understanding of the genes involved in antibiotic resistance in Mycobacterium tuberculosis (Mtb) is necessary for the design of improved therapies. Clustered regularly interspaced short palindromic repeat interference (CRISPRi) has been previously utilized in mycobacteria to identify novel drug targets by the demonstration of gene essentiality. The work presented here shows that it can also be usefully applied to the study of non-essential genes involved in antibiotic resistance. The expression of an ADP-ribosyltransferase (Arr) involved in rifampicin resistance in Mycobacterium smegmatis was silenced using CRISPRi and the impact on rifampicin susceptibility was measured. Gene silencing resulted in a decrease in the minimum inhibitory concentration (MIC) similar to that previously reported in an arr deletion mutant. There is contradictory evidence for the toxicity of Streptococcus pyogenes dCas9 (dCas9 Spy) in the literature. In this study the expression of dCas9 Spy in M. smegmatis showed no impact on viability. Silencing was achieved with concentrations of the aTc inducer lower than previously described and with shorter induction times. Finally, designing small guide RNAs (sgRNAs) that target transcription initiation, or the early stages of elongation had the most impact on rifampicin susceptibility. This study demonstrates that CRISPRi based gene silencing can be as impactful as gene deletion for the study of non-essential genes and further contributes to the knowledge on the design and induction of sgRNAs for CRISPRi. This approach can be applied to other non-essential antimicrobial resistance genes such as drug efflux pumps.Peer reviewe
Towards an OpenSource Logger for the Analysis of RPA Projects
Process automation typically begins with the observation of
humans conducting the tasks that will be eventually automated. Sim ilarly, successful RPA projects require a prior analysis of the undergo ing processes which are being executed by humans. The process of col lecting this type of information is known as user interface (UI) logging
since it records the interaction against a UI. Main RPA platforms (e.g.,
Blueprism and UIPath) incorporate functionalities that allow the record ing of these UI interactions. However, the records that these platforms
generate lack some functionalities that large-scale RPA projects require.
Besides, they are only understandable by the proper RPA platforms.
This paper presents an extensible and multi-platform OpenSource UI
logger that generate UI logs in a standard format. This system collects
information from all the computers it is running on and sends it to a
central server for its processing. Treatment of the collected information
will allow the creation of an enriched UI log which can be used, among
others purposes, for smart process analysis, machine learning training,
the creation of RPA robots, or, being more general, for task mining .Ministerio de EconomÃa y Competitividad TIN2016-76956-C3-2-R (POLOLAS)Junta de AndalucÃa CEI-12-TIC021Centro para el Desarrollo Tecnol´ogico Industrial (CDTI) P011-19/E0
Inactivation of bpsl1039-1040 ATP-binding cassette transporter reduces intracellular survival in macrophages, biofilm formation and virulence in the murine model of Burkholderia pseudomallei infection.
Burkholderia pseudomallei, a gram-negative intracellular bacillus, is the causative agent of a tropical infectious disease called melioidosis. Bacterial ATP-binding cassette (ABC) transporters import and export a variety of molecules across bacterial cell membranes. At present, their significance in B. pseudomallei pathogenesis is poorly understood. We report here characterization of the BPSL1039-1040 ABC transporter. B. pseudomallei cultured in M9 medium supplemented with nitrate, demonstrated that BPSL1039-1040 is involved in nitrate transport for B. pseudomallei growth under anaerobic, but not aerobic conditions, suggesting that BPSL1039-1040 is functional under reduced oxygen tension. In addition, a nitrate reduction assay supported the function of BPSL1039-1040 as nitrate importer. A bpsl1039-1040 deficient mutant showed reduced biofilm formation as compared with the wild-type strain (P = 0.027) when cultured in LB medium supplemented with nitrate under anaerobic growth conditions. This reduction was not noticeable under aerobic conditions. This suggests that a gradient in oxygen levels could regulate the function of BPSL1039-1040 in B. pseudomallei nitrate metabolism. Furthermore, the B. pseudomallei bpsl1039-1040 mutant had a pronounced effect on plaque formation (P < 0.001), and was defective in intracellular survival in both non-phagocytic (HeLa) and phagocytic (J774A.1 macrophage) cells, suggesting reduced virulence in the mutant strain. The bpsl1039-1040 mutant was found to be attenuated in a BALB/c mouse intranasal infection model. Complementation of the bpsl1039-1040 deficient mutant with the plasmid-borne bpsl1039 gene could restore the phenotypes observed. We propose that the ability to acquire nitrate for survival under anaerobic conditions may, at least in part, be important for intracellular survival and has a contributory role in the pathogenesis of B. pseudomallei
UK Housing Market: Time Series Processes with Independent and Identically Distributed Residuals
The paper examines whether a univariate data generating process can be identified which explains the data by having residuals that are independent and identically distributed, as verified by the BDS test. The stationary first differenced natural log quarterly house price index is regressed, initially with a constant variance and then with a conditional variance. The only regression function that produces independent and identically distributed standardised residuals is a mean process based on a pure random walk format with Exponential GARCH in mean for the conditional variance. There is an indication of an asymmetric volatility feedback effect but higher frequency data is required to confirm this. There could be scope for forecasting the index but this is tempered by the reduction in the power of the BDS test if there is a non-linear conditional variance process
- …