Pseudolysogeny and sequential mutations build multiresistance to virulent bacteriophages in Pseudomonas aeruginosa

International audienceCoevolution between bacteriophages and their prey is the result of mutualistic interactions. Here we show that pseudolysogeny is a frequent outcome of infection by virulent phages of Pseudomonas aeruginosa, and that selection of resistant bacterial mutants is favored by continuous production of phages. We investigated the frequency and characteristics of P. aeruginosa strain PAO1 variants resisting infection by different combinations of virulent phages belonging to four genera. The frequency of resistant bacteria was 10-5 for single phage infection and 10-6 for infections with combinations of two or four phages. The genome of 27 variants was sequenced and the comparison with the genome of the parental PAO1 strain allowed the identification of point mutations or small indels. Four additional variants were characterized by a candidate gene approach. In total, 27 independent mutations were observed affecting 14 genes and a regulatory region. The mutations affected genes involved in biosynthesis of type IV pilus, alginate, LPS and O-antigen. Half of the variants possessed changes in homopolymer tracts responsible for frameshift mutations, and these phase variation mutants were shown to be unstable. Eleven double mutants were detected. The presence of free phage DNA was observed in association with exclusion of superinfection in half of the variants, and in three of them no chromosomal mutation could be found. Upon further growth of these pseudolysogens, some variants with new chromosomal mutations were recovered presumably due to continuous evolutionary pressure

Relativism in the linguistic representation and cognitive conceptualisation of motion events across verb-framed and satellite-framed languages

The present doctoral thesis addresses the issue of the relation in human cognition between language and thinking, and, more specifically, it aims to investigate by scientific means the potential for a language-particular influence on cognitive activity and putative reflexes, i.e. the linguistic relativity question (cf. Whorf 1956, Lucy 1992a).To this end, the present thesis offers a detailed exploration of linguistic relativity and of its potential scope of validity - at least in theoretical terms. It further situates its study within modern cognitive science, whose epistemological approach to the study of the mind is multi- disciplinary, bringing the fields of psychology, linguistics and philosophy together for the enhanced pursuit of an understanding of human cognition. Having established a conducive framework for the study of linguistic relativity within cognitive science and linguistics, the thesis offers to focus on a specific experiential domain of human life, and on its variable encoding in different languages to seek specific language influences over the conceptualisation of that domain. The chosen domain consists of MOTION - a pervasive domain in humans' daily lives and daily needs of expression. This domain is particularly interesting to relativistic studies as its conceptual components are lexicalised via differing means across the world's languages. Existing typologies for motion encoding (e.g. Talmy 1985) have established at least two main possible patterns, also known as verb- and satellite-framing, and as exemplified by the French and English languages respectively. The essential difference between the two language types consists of their grammatical encoding of the core element of motion, namely PATH - either in a verb or in a verbal satellite ― and of their selective encoding of peripheral elements, such as MANNER of displacement - with this element being optional in French grammar, and obligatory in English. The thesis offers empirical linguistic data to confirm - and also challenge - the fixedness of the patterns identified by e.g. Talmy. A thorough discussion of the linguistic framing of motion is presented, together with experiments bearing on the cognitive reality of motion conceptualisation - independently of language. This thesis thus contributes to an understanding of motion both in language and in cognition. Finally, it offers experimental work bearing on the relativity question, i.e. exploring whether linguistic patterns for motion encoding exert a decisive influence on the non-linguistic conceptualisation of motion, resulting in the two language communities differing in their cognitive appreciation of otherwise similar motion events. The final results offer evidence in favour of differing conceptualisations, that is, in support of linguistic relativity

Bio-coloration of bacterial cellulose assisted by immobilized laccase

In this work a process for the bio-coloration of bacterial cellulose (BC) membranes was developed. Laccase from Myceliophthora thermophila was immobilized onto BC membranes and retained up to 88% of residual activity after immobilization. Four compounds belonging to the flavonoids family were chosen to test the in situ polymerase activity of immobilized laccase. All the flavonoids were successfully polymerized by laccase giving rise to yellow, orange and dark brown oligomers which conferred color to the BC support. The optimal bio-coloration conditions were studied for two of the tested flavonoids, catechol and catechin, by varying the concentration and time of incubation. High color depth and resistance to washing were obtained for both compounds. The highly porous bacterial cellulose material demonstrated great performance as a bio-coloration support, in contrast to other materials cited in literature, like cotton or wool. The process developed is presented as an environmentally friendly alternative for bacterial cellulose bio-coloration and will contribute deeply for the development of new fashionable products within this material.The authors would like to acknowledge Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI‑01‑0145‑FEDER‑006684) and BioTecNorte operation (NORTE‑01‑0145‑FEDER‑000004) funded by Euro‑ pean Regional Development Fund under the scope of Norte2020‑Programa Operacional Regional do Norte. The authors would like also to acknowl‑ edge the Basic Science Research Program through the National Research Foundation of Korea (NRF), which was funded by the Ministry of Education (2017R1D1A1B03031959).info:eu-repo/semantics/publishedVersio

How the other half lives: CRISPR-Cas's influence on bacteriophages

CRISPR-Cas is a genetic adaptive immune system unique to prokaryotic cells used to combat phage and plasmid threats. The host cell adapts by incorporating DNA sequences from invading phages or plasmids into its CRISPR locus as spacers. These spacers are expressed as mobile surveillance RNAs that direct CRISPR-associated (Cas) proteins to protect against subsequent attack by the same phages or plasmids. The threat from mobile genetic elements inevitably shapes the CRISPR loci of archaea and bacteria, and simultaneously the CRISPR-Cas immune system drives evolution of these invaders. Here we highlight our recent work, as well as that of others, that seeks to understand phage mechanisms of CRISPR-Cas evasion and conditions for population coexistence of phages with CRISPR-protected prokaryotes.Comment: 24 pages, 8 figure

CRISPRFinder: a web tool to identify clustered regularly interspaced short palindromic repeats

Clustered regularly interspaced short palindromic repeats (CRISPRs) constitute a particular family of tandem repeats found in a wide range of prokaryotic genomes (half of eubacteria and almost all archaea). They consist of a succession of highly conserved regions (DR) varying in size from 23 to 47 bp, separated by similarly sized unique sequences (spacer) of usually viral origin. A CRISPR cluster is flanked on one side by an AT-rich sequence called the leader and assumed to be a transcriptional promoter. Recent studies suggest that this structure represents a putative RNA-interference-based immune system. Here we describe CRISPRFinder, a web service offering tools to (i) detect CRISPRs including the shortest ones (one or two motifs); (ii) define DRs and extract spacers; (iii) get the flanking sequences to determine the leader; (iv) blast spacers against Genbank database and (v) check if the DR is found elsewhere in prokaryotic sequenced genomes. CRISPRFinder is freely accessible at http://crispr.u-psud.fr/Server/CRISPRfinder.php

The CRISPRdb database and tools to display CRISPRs and to generate dictionaries of spacers and repeats

<p>Abstract</p> <p>Background</p> <p>In Archeae and Bacteria, the repeated elements called CRISPRs for "clustered regularly interspaced short palindromic repeats" are believed to participate in the defence against viruses. Short sequences called spacers are stored in-between repeated elements. In the current model, motifs comprising spacers and repeats may target an invading DNA and lead to its degradation through a proposed mechanism similar to RNA interference. Analysis of intra-species polymorphism shows that new motifs (one spacer and one repeated element) are added in a polarised fashion. Although their principal characteristics have been described, a lot remains to be discovered on the way CRISPRs are created and evolve. As new genome sequences become available it appears necessary to develop automated scanning tools to make available CRISPRs related information and to facilitate additional investigations.</p> <p>Description</p> <p>We have produced a program, CRISPRFinder, which identifies CRISPRs and extracts the repeated and unique sequences. Using this software, a database is constructed which is automatically updated monthly from newly released genome sequences. Additional tools were created to allow the alignment of flanking sequences in search for similarities between different loci and to build dictionaries of unique sequences. To date, almost six hundred CRISPRs have been identified in 475 published genomes. Two Archeae out of thirty-seven and about half of Bacteria do not possess a CRISPR. Fine analysis of repeated sequences strongly supports the current view that new motifs are added at one end of the CRISPR adjacent to the putative promoter.</p> <p>Conclusion</p> <p>It is hoped that availability of a public database, regularly updated and which can be queried on the web will help in further dissecting and understanding CRISPR structure and flanking sequences evolution. Subsequent analyses of the intra-species CRISPR polymorphism will be facilitated by CRISPRFinder and the dictionary creator. CRISPRdb is accessible at <url>http://crispr.u-psud.fr/crispr</url></p

Management et performances d'un processus de formation

Cet article a pour objectif de présenter une approche à  la conception de la stratégie des systêmes de production de compétences. Cette méthode est inspirée des approches actuelles dans de nombreux secteurs des systêmes de production de biens et de services. L'approche proposée conduit à  disposer d'un modêle du systême étudié, d'une expression structurée de sa stratégie avant de définir et de la déployer selon une phase structurée de conception du pilotage. L'article montre qu'il est intéressant de définir, dans un premier temps, les processus significatifs afin de limiter la démarche de déploiement de la stratégie

Rethinking 'Thinking for Speaking'

Proceedings of the Twenty-Ninth Annual Meeting of the Berkeley Linguistics Society: General Session and Parasession on Phonetic Sources of Phonological Patterns: Synchronic and Diachronic Explanations (2003

Structural basis for CRISPR RNA-guided DNA recognition by Cascade

The CRISPR (clustered regularly interspaced short palindromic repeats) immune system in prokaryotes uses small guide RNAs to neutralize invading viruses and plasmids. In Escherichia coli, immunity depends on a ribonucleoprotein complex called Cascade. Here we present the composition and low-resolution structure of Cascade and show how it recognizes double-stranded DNA (dsDNA) targets in a sequence-specific manner. Cascade is a 405-kDa complex comprising five functionally essential CRISPR-associated (Cas) proteins (CasA1B2C6D1E1) and a 61-nucleotide CRISPR RNA (crRNA) with 5′-hydroxyl and 2′,3′-cyclic phosphate termini. The crRNA guides Cascade to dsDNA target sequences by forming base pairs with the complementary DNA strand while displacing the noncomplementary strand to form an R-loop. Cascade recognizes target DNA without consuming ATP, which suggests that continuous invader DNA surveillance takes place without energy investment. The structure of Cascade shows an unusual seahorse shape that undergoes conformational changes when it binds target DNA.