Contribution of teg49 Small RNA in the 5′ Upstream Transcriptional Region of sarA to Virulence in Staphylococcus aureus

High-throughput RNA sequencing technology has found the 5\u27 untranslated region of sarA to contain two putative small RNAs (sRNAs), designated teg49 and teg48. Northern blot analysis disclosed that teg49 and teg48 were detectable within the P3-P1 and P1 sarA promoter regions, respectively. Focusing on teg49, we found that this sRNA, consisting of 196 nucleotides, is transcribed in the same direction as the sarA P3 transcript. The expression of both P3 and teg49 transcripts is dependent on sigB and cshA, which encodes a DEAD box RNA helicase. Within the sRNA teg49, there are two putative hairpin-loop structures, HP1 and HP2. Transversion mutation of the HP1 loop produced a smaller amount of sarA P3 and P2 transcripts and SarA protein than the corresponding HP1 stem and the HP2 stem and loop mutations, leading to lower RNAII transcription and derepression of aur transcription. The HP1 loop mutant also exhibited less biofilm formation than the parental and complemented strains. Complementation with shuttle plasmid pEPSA5 carrying teg49 was able to reestablish sarA P3 and P2 transcription and augment RNAII expression in the HP1 loop mutant. We thus conclude that teg49, embedded within the extended promoter regions of sarA, is modulated by sigB and cshA and plays an important trans-acting role in modulating the transcription and ensuing expression of sarA

Contribution of teg49 Small RNA in the 5′ Upstream Transcriptional Region of sarA to Virulence in Staphylococcus aureus

High-throughput RNA sequencing technology has found the 5\u27 untranslated region of sarA to contain two putative small RNAs (sRNAs), designated teg49 and teg48. Northern blot analysis disclosed that teg49 and teg48 were detectable within the P3-P1 and P1 sarA promoter regions, respectively. Focusing on teg49, we found that this sRNA, consisting of 196 nucleotides, is transcribed in the same direction as the sarA P3 transcript. The expression of both P3 and teg49 transcripts is dependent on sigB and cshA, which encodes a DEAD box RNA helicase. Within the sRNA teg49, there are two putative hairpin-loop structures, HP1 and HP2. Transversion mutation of the HP1 loop produced a smaller amount of sarA P3 and P2 transcripts and SarA protein than the corresponding HP1 stem and the HP2 stem and loop mutations, leading to lower RNAII transcription and derepression of aur transcription. The HP1 loop mutant also exhibited less biofilm formation than the parental and complemented strains. Complementation with shuttle plasmid pEPSA5 carrying teg49 was able to reestablish sarA P3 and P2 transcription and augment RNAII expression in the HP1 loop mutant. We thus conclude that teg49, embedded within the extended promoter regions of sarA, is modulated by sigB and cshA and plays an important trans-acting role in modulating the transcription and ensuing expression of sarA

Microbial Communities of Conducting and Respiratory Zones of Lung-Transplanted Patients.

Background: Lung transplantation (LT) is a recognized treatment for end-stage pulmonary disease. Bacteria from the recipient nasopharynx seed the new lungs leading to infections and allograft damage. Understanding the characteristics and topological variations of the microbiota may be important to apprehend the pathophysiology of allograft dysfunction. Objectives: To examine the characteristics and relationship of bacterial compositions between conducting and respiratory zones of the allograft. Methods: We performed 16S rRNA gene sequencing on bronchial aspirates (BAs) and bronchoalveolar lavages (BALs) collected in pairs in 19 patients at several time-points post-LT. Results: The respiratory zone was characterized independently of the time post-LT by a higher bacterial richness than the conducting zone (p = 0.041). The phyla Firmicutes and Proteobacteria dominated both sampling zones, with an inverse correlation between these two phyla (Spearman r = -0.830). Samples of the same pair, as well as pairs from the same individual clustered together (Pseudo-F = 3.8652, p < 0.01). Microbiota of BA and BAL were more closely related in samples from the same patient than each sample type across different patients, with variation in community structure being mainly inter-individual (p < 0.01). Both number of antibiotics administered (p < 0.01) and time interval post-LT (p < 0.01) contributed to the variation in global microbiota structure. Longitudinal analysis of BA-BAL pairs of two patients showed dynamic wave like fluctuations of the microbiota. Conclusions: Our results show that post-transplant respiratory zones harbor higher bacterial richness, but overall similar bacterial profiles as compared to conductive zones. They further support an individual microbial signature following LT

Hot tearing test for TIG welding of aluminum alloys: application of a stress parallel to the fusion line

Welding processes are extensively used to assemble components in many manufacturing industries, such as aeronautics, construction, energy and automotive. To increase their productivity, constructors try to reduce manufacturing time. This involves for welding operations an increase of welding speed. However, various defects such as hot tearing might limit this increase. Hot tearing appears at the solidification end of an alloy, and is commonly observed in welding. During welding, components are subjected to high thermal gradients around the melting zone due to localized heat input. The solidification zone, where microstructure forms, is located at the rear of the melting zone and is bordered by two isothermal surfaces corresponding to liquidus and solidus temperatures. In welding, due to high thermal gradients, solidification by epitaxial growth of columnar dendrites is generally observed from the border of the melting zone. However, equiaxed dendritic grains can also form in the center area of the melting zone

Cartography of Methicillin-Resistant S. aureus Transcripts: Detection, Orientation and Temporal Expression during Growth Phase and Stress Conditions

BACKGROUND: Staphylococcus aureus is a versatile bacterial opportunist responsible for a wide spectrum of infections. The severity of these infections is highly variable and depends on multiple parameters including the genome content of the bacterium as well as the condition of the infected host. Clinically and epidemiologically, S. aureus shows a particular capacity to survive and adapt to drastic environmental changes including the presence of numerous antimicrobial agents. Mechanisms triggering this adaptation remain largely unknown despite important research efforts. Most studies evaluating gene content have so far neglected to analyze the so-called intergenic regions as well as potential antisense RNA molecules. PRINCIPAL FINDINGS: Using high-throughput sequencing technology, we performed an inventory of the whole transcriptome of S. aureus strain N315. In addition to the annotated transcription units, we identified more than 195 small transcribed regions, in the chromosome and the plasmid of S. aureus strain N315. The coding strand of each transcript was identified and structural analysis enabled classification of all discovered transcripts. RNA purified at four time-points during the growth phase of the bacterium allowed us to define the temporal expression of such transcripts. A selection of 26 transcripts of interest dispersed along the intergenic regions was assessed for expression changes in the presence of various stress conditions including pH, temperature, oxidative shocks and growth in a stringent medium. Most of these transcripts showed expression patterns specific for the defined stress conditions that we tested. CONCLUSIONS: These RNA molecules potentially represent important effectors of S. aureus adaptation and more generally could support some of the epidemiological characteristics of the bacterium

Detection and characterization of small RNAs in Staphylococcus aureus

Staphylococcus aureus est un pathogène majeur causant une large diversité d'infections. Cette bactérie démontre une grande capacité à d'adaptation, suggérant une fine régulation de l'expression des gènes. Les petits ARNs sont reconnus comme des contributeurs majeurs de la régulation transcriptionnelle. Chez S. aureus, très peu de petits ARNs restaient cependant identifiés et caractérisés. Ce travail a permis de montrer que, chez S. aureus, plusieurs centaines de petits ARNs étaient transcrits à partir des régions intergéniques et en antisens à des gènes annotés. Ces nouveaux transcrits ont été classifiés en 8 sous-catégories d'ARN régulateurs. De plus, de vastes études d'expression ont démontré que la transcription de ces molécules était elle-même largement soumise à régulation. Le rôle de 8 de ces transcrits dans la biologie du pathogène a été investigué. Teg131 pourrait être un régulateur clé de l'expression de gènes associés à la virulence et/ou au métabolisme de biosynthèse de la paroi

New approaches for functional genomic studies in staphylococci

Functional transcriptomics studies have resulted in interesting insights into Staphylococcus aureus diversity and pathogenicity. Here we review the principles, advantages and disadvantages of recent technical developments in the field of transcriptomics and their potential impact on S. aureus research

Bacterial genome evolution within a clonal population: from in vitro investigations to in vivo observations

Bacteria are faced with a diversity of environmental stresses that include high salt concentrations, heavy metals and pH fluctuations. Adaptation to resist such stresses is a complex phenomenon that involves global pathways and simultaneous acquisition of multiple unrelated properties. During the last 3 years, the development of new technologies in the field of molecular biology has led to numerous fundamental and quantitative in vitro and in vivo evolutionary studies that have improved our understanding of the principles underlying bacterial adaptations, and helped us develop strategies to cope with the health burden of bacterial virulence. In this review, the authors discuss the evolution of bacteria in the laboratory and in human patients

Hot tearing test for TIG welding of aluminium alloys: application of a tensile load parallel to the fusion line

International audienceDefects control such as hot cracking in aluminum alloys welding is an important industrial issue. Understanding of hot cracking phenomenon is a complex problem involving process, material and mechanical loading due to clamping. Several tests have been previously developed in order to characterize the material propensity to hot cracking. The purpose of the present work is to study, using a new hot cracking test and numerical simulation, the relationship between mechanical and metallurgical factors in order to better identify the parameters leading to hot tearing during welding. The originality of the test presented here is that an external stress is applied on the test specimen parallel to the welding direction. The advantage of this test, compared to others like Varestraint test, is its simplicity which is interesting for an industrial use. A fusion line is made with a Tungsten Inert Gas (TIG) arc welding process on a thin sheet of aluminum alloy (6061). The crack initiation occurs once steady state thermal conditions are reached. The present test enables to distinguish between the structural effects on a global scale and the microstructural effects on a local scale. Microstructure control is made possible by adjusting welding power, welding speed and sample geometries. The grain morphology plays a crucial role in the crack initiation. It depends on the intensity and the welding speed and the specimen geometry. It is characterized by the shape, size but also the direction of grain growth that influence the cracks initiation. Microstructural features are observed using high speed camera recording and post mortem micrographs. Mechanical factors are varied by adjusting the welding parameters and the applied prestress. The relationship between welding parameters, morphology of the generated grain structure, and sensitivity to hot cracking were discussed. Experimental measurements and numerical results will help to better determine global and local conditions at the onset of hot tearing and to compare those conditions using existing hot tearing criteria