The T7-Related Pseudomonas putida Phage ϕ15 Displays Virion-Associated Biofilm Degradation Properties

Formation of a protected biofilm environment is recognized as one of the major causes of the increasing antibiotic resistance development and emphasizes the need to develop alternative antibacterial strategies, like phage therapy. This study investigates the in vitro degradation of single-species Pseudomonas putida biofilms, PpG1 and RD5PR2, by the novel phage ϕ15, a ‘T7-like virus’ with a virion-associated exopolysaccharide (EPS) depolymerase. Phage ϕ15 forms plaques surrounded by growing opaque halo zones, indicative for EPS degradation, on seven out of 53 P. putida strains. The absence of haloes on infection resistant strains suggests that the EPS probably act as a primary bacterial receptor for phage infection. Independent of bacterial strain or biofilm age, a time and dose dependent response of ϕ15-mediated biofilm degradation was observed with generally a maximum biofilm degradation 8 h after addition of the higher phage doses (104 and 106 pfu) and resistance development after 24 h. Biofilm age, an in vivo very variable parameter, reduced markedly phage-mediated degradation of PpG1 biofilms, while degradation of RD5PR2 biofilms and ϕ15 amplification were unaffected. Killing of the planktonic culture occurred in parallel with but was always more pronounced than biofilm degradation, accentuating the need for evaluating phages for therapeutic purposes in biofilm conditions. EPS degrading activity of recombinantly expressed viral tail spike was confirmed by capsule staining. These data suggests that the addition of high initial titers of specifically selected phages with a proper EPS depolymerase are crucial criteria in the development of phage therapy

Purification and determination of the structure of capsular polysaccharide of Vibrio vulnificus M06-24.

Virulence of Vibrio vulnificus has been strongly associated with encapsulation and an opaque colony morphology. Capsular polysaccharide was purified from a whole-cell, phosphate-buffered saline-extracted preparation of the opaque, virulent phase of V. vulnificus M06-24 (M06-24/O) by dialysis, centrifugation, enzymatic digestion, and phenol-chloroform extraction. Nuclear magnetic resonance spectroscopic analysis of the purified polysaccharide showed that the polymer was composed of a repeating structure with four sugar residues per repeating subunit: three residues of 2-acetamido-2,6-dideoxyhexopyranose in the alpha-gluco configuration (QuiNAc) and an additional residue of 2-acetamido hexouronate in the alpha-galactopyranose configuration (GalNAcA). The complete carbohydrate structure of the polysaccharide was determined by heteronuclear nuclear magnetic resonance spectroscopy and by high-performance anion-exchange chromatography. The 1H and 13C nuclear magnetic resonance spectra were completely assigned, and vicinal coupling relationships were used to establish the stereochemistry of each sugar residue, its anomeric configuration, and the positions of the glycosidic linkages. The complete structure is: [----3) QuipNAc alpha-(1----3)-GalpNAcA alpha-(1----3)-QuipNAc alpha-(1----]n QuipNAc alpha-(1----4)-increases The polysaccharide was produced by a translucent phase variant of M06-24 (M06-24/T) but not by a translucent, acapsular transposon mutant (CVD752). Antibodies to the polysaccharide were demonstrable in serum from rabbits inoculated with M06-24/O

An alternating algorithm for Prompt Gamma Time Imaging with the TIARA project

International audienceIn hadrontherapy, the precise tracking of the hadron during treatment and the real-time determination of its different characteristics (prompt gamma positions, velocity profile and energy loss) is highly recommended to detect any change or mismatch between the treatment planning and the conditions present at the time of the treatment.To the best of our knowledge, there is currently no automatic method proposed to meet this need and promptly alert or halt the treatment if there is a risk of over-irradiating healthy tissues.In the present work, we present a mathematical measurement model and an alternating strategy that ensure a joint estimation of the different properties of the hadron (the prompt gamma positions and the velocity profile allow us to determine the energy loss) throughout the measured times of flight. Our proposition is based on convergence guarantees

An alternating algorithm for Prompt Gamma Time Imaging with the TIARA project

International audienceIn hadrontherapy, the precise tracking of the hadron during treatment and the real-time determination of its different characteristics (prompt gamma positions, velocity profile and energy loss) is highly recommended to detect any change or mismatch between the treatment planning and the conditions present at the time of the treatment.To the best of our knowledge, there is currently no automatic method proposed to meet this need and promptly alert or halt the treatment if there is a risk of over-irradiating healthy tissues.In the present work, we present a mathematical measurement model and an alternating strategy that ensure a joint estimation of the different properties of the hadron (the prompt gamma positions and the velocity profile allow us to determine the energy loss) throughout the measured times of flight. Our proposition is based on convergence guarantees