Detection of single DNA molecules by multicolor quantum-dot end-labeling

Observation of DNA–protein interactions by single molecule fluorescence microscopy is usually performed by using fluorescent DNA binding agents. However, such dyes have been shown to induce cleavage of the DNA molecule and perturb its interactions with proteins. A new method for the detection of surface-attached DNA molecules by fluorescence microscopy is introduced in this paper. Biotin- and/or digoxigenin-modified DNA fragments are covalently linked at both extremities of a DNA molecule via sequence-specific hybridization and ligation. After the modified DNA molecules have been stretched on a glass surface, their ends are visualized by multicolor fluorescence microscopy using conjugated quantum dots (QD). We demonstrate that under carefully selected conditions, the position and orientation of individual DNA molecules can be inferred with good efficiency from the QD fluorescence signals alone. This is achieved by selecting QD pairs that have the distance and direction expected for the combed DNA molecules. Direct observation of single DNA molecules in the absence of DNA staining agent opens new possibilities in the fundamental study of DNA–protein interactions. This work also documents new possibilities regarding the use of QD for nucleic acid detection and analysis

Comparative efficacy of two fipronil spot-on formulations against experimental tick infestations (Ixodes ricinus) in dogs

A parallel-group-design, randomized, unicentre and blinded controlled study was undertaken to assess the efficacy of a new fipronil-based spot-on formulation applied once to dogs against experimental Ixodes ricinus infestations. Six dogs served as negative controls (group 1), six dogs served as positive controls (group 2) receiving the original fipronil spot-on (Frontline® spot-on Dog, Merial) at a dosage of 0.67 mL for a dog weighing from 2 to 10 kg and 1.34 mL for a dog weighing from 10.1 to 20 kg and six dogs were treated with a 10% w/v fipronil-based spot-on solution (Effipro® Spot-on, Virbac SA) at an identical dosage (group 3, 0.67 mL for a dog weighing from 2 to 10 kg and 1.34 mL for a dog weighing from 10.1 to 20 kg). Each dog was sedated and subsequently infested with 50 unfed adult I. ricinus on days −7, −2, 7, 14, 21, 28 and 35. Forty-eight hours after the treatment and 48 h after each challenge (days −5, 2, 9, 16, 23, 30 and 37), the population of the remaining ticks was assessed for each animal. Geometric mean tick counts obtained were reduced by 99% and 94% on day 2 in groups 2 and 3, respectively, compared to the negative control group. Dogs were protected from re-infestations with an efficacy of >90% for 3 weeks in group 2 and for 5 weeks in group 3. Both 10% w/v fipronil-based spot-on solutions, despite different vehicles, were equally able to eradicate tick infestation, to prevent new infestations and were equally well tolerated

New evidence of a mitochondrial genetic background paradox: Impact of the J haplogroup on the A3243G mutation

International audienceBackground: The A3243G mutation in the tRNALeu gene (UUR), is one of the most common pathogenic mitochondrial DNA (mtDNA) mutations in France, and is associated with highly variable and heterogeneous disease phenotypes. To define the relationships between the A3243G mutation and mtDNA backgrounds, we determined the haplogroup affiliation of 142 unrelated French patients – diagnosed as carriers of the A3243G mutation – by control-region sequencing and RFLP survey of their mtDNAs. Results: The analysis revealed 111 different haplotypes encompassing all European haplogroups, indicating that the 3243 site might be a mutational hot spot. However, contrary to previous findings, we observed a statistically significant underepresentation of the A3243G mutation on haplogroup J in patients (p = 0.01, OR = 0.26, C.I. 95%: 0.08–0.83), suggesting that might be due to a strong negative selection at the embryo or germ line stages. Conclusion: Thus, our study supports the existence of mutational hotspot on mtDNA and a "haplogroup J paradox," a haplogroup that may increase the expression of mtDNA pathogenic mutations, but also be beneficial in certain environmental contexts

Genome sequencing of Xanthomonas axonopodis pv. phaseoli CFBP4834-R reveals that flagellar motility is not a general feature of xanthomonads.

Xanthomonads are plant-associated bacteria that establish neutral, commensal or pathogenic relationships with plants. The list of common characteristics shared by all members of the genus Xanthomonas is now well established based on the entire genome sequences that are currently available and that represent various species, numerous pathovars of X. axonopodis (sensu Vauterin et al., 2000), X. oryzae and X. campestris, and many strains within some pathovars. These ?-proteobacteria are motile by a single polar flagellum. Motility is an important feature involved in biofilm formation, plant colonization and hence considered as a pathogenicity factor. X. axonopodis pv. phaseoli var. fuscans (Xapf) is one of the causal agents of common bacterial blight of bean and 4834-R is a highly aggressive strain of this pathogen that was isolated from a seed-borne epidemic in France in 1998. We obtained a high quality assembled sequence of the genome of this strain with 454-Solexa and 2X Sanger sequencing. Housekeeping functions are conserved in this genome that shares core characteristics with genomes of other xanthomonads: the six secretion systems which have been described so far in Gram negative bacteria are all present, as well as their ubiquitous substrates or effectors and a rather usual number of mobile elements. Elements devoted to the adaptation to the environment constitute an important part of the genome with a chemotaxis island and dispersed MCPs, numerous two-component systems, and numerous TonB dependent transporters. Furthermore, numerous multidrug efflux systems and functions dedicated to biofilm formation that confer resistance to stresses are also present. An intriguing feature revealed by genome analysis is a long deletion of 35 genes (33 kbp) involved in flagellar biosynthesis. This deletion is replaced by an insertion sequence called ISXapf2. Genes such as flgB to flgL and fliC to fleQ which are involved in the flagellar structure (rod, P- and L-ring, hook, cap and filament) are absent in the genome of strain 4834-R that is not motile. Primers were designed to detect this deletion by PCR in a collection of more than 300 strains representing different species and pathovars of Xanthomonas, and less than 5% of the tested xanthomonads strains were found nonmotile because of a deletion in the flagellum gene cluster. We observed that half of the Xapf strains isolated from the same epidemic than strain 4834-R was non-motile and that this ratio was conserved in the strains colonizing the next bean seed generation. Isolation of such variants in a natural epidemic reveals that either flagellar motility is not a key function for fitness or that some complementation occurs within the bacterial population. (Résumé d'auteur

Genome sequence of Xanthomonas fuscans subsp. fuscans strain 4834-R reveals that flagellar motility is not a general feature of xanthomonads

Abstract\ud \ud \ud \ud Background\ud Xanthomonads are plant-associated bacteria responsible for diseases on economically important crops. Xanthomonas fuscans subsp. fuscans (Xff) is one of the causal agents of common bacterial blight of bean. In this study, the complete genome sequence of strain Xff 4834-R was determined and compared to other Xanthomonas genome sequences.\ud \ud \ud \ud Results\ud Comparative genomics analyses revealed core characteristics shared between Xff 4834-R and other xanthomonads including chemotaxis elements, two-component systems, TonB-dependent transporters, secretion systems (from T1SS to T6SS) and multiple effectors. For instance a repertoire of 29 Type 3 Effectors (T3Es) with two Transcription Activator-Like Effectors was predicted. Mobile elements were associated with major modifications in the genome structure and gene content in comparison to other Xanthomonas genomes. Notably, a deletion of 33 kbp affects flagellum biosynthesis in Xff 4834-R. The presence of a complete flagellar cluster was assessed in a collection of more than 300 strains representing different species and pathovars of Xanthomonas. Five percent of the tested strains presented a deletion in the flagellar cluster and were non-motile. Moreover, half of the Xff strains isolated from the same epidemic than 4834-R was non-motile and this ratio was conserved in the strains colonizing the next bean seed generations.\ud \ud \ud \ud Conclusions\ud This work describes the first genome of a Xanthomonas strain pathogenic on bean and reports the existence of non-motile xanthomonads belonging to different species and pathovars. Isolation of such Xff variants from a natural epidemic may suggest that flagellar motility is not a key function for in planta fitness.AI is funded by a PhD grant from INRA-SPE and region Pays de la Loire, France. EG was funded by a PhD grant from the French Ministry of National Education and Research and French Guyana. SC, EG, MA, EL and LDN are funded by the LABEX TULIP (ANR-10-LABX-41), LSG is funded by ANR-2010-GENM-013 Xanthomix

Saft VES16 Solution for Small GEO

At the end of 2008, Saft has designed the VES16 cell [1], a new Li-ion cell aiming to address Low Earth Orbit (LEO) satellite market. In the frame of ESA GSTP 5.2 activity, Saft has then developed and successfully qualified [2] a range of batteries based on the VES16 cell and its autonomous Simplified Balancing System (SBS), targeting both LEO and Geostationary Earth Orbit (GEO) missions The very good performance of the VES16 cell demonstrated in GEO mission cycling configuration allows now Saft to propose it on small geostationary spacecraft with typical power lower than 10kW, offering a simpler, autonomous and competitive solution compared to Saft high capacity cells solutions with VES180 qualified on several telecommunication spacecraft. This paper will present the design of the VES16, its performance for GEO mission, the configuration of batteries based on this cell, with their benefit and drawbacks, and will detail two current GEO applications using VES16 batteries

A framework towards personalisation and active muscle integration in a 3D finite-element neck model for orthopaedic applications

39e Congrès de la Société de Biomécanique, VALENCIENNES, FRANCE, 27-/08/2014 - 29/08/2014In biomechanics, multi-segmented rigid-body and finite element (FE) models have become an essential tool to explore and quantify the dynamics of human motion through objective parameters. At the neck level, numerous segmental and parametric FE approaches have been used to quantify loads (e.g. to evaluate the design of spinal devices), but only a few studies have explored the influence of subject-specific geometric postural variations on the cervical spine's behaviour (Frechede et al. 2006; Laville et al. 2009). Explorations of the influence of the muscles and the subject-specific complexity of their load sharing on the dynamic stability of the cervical spine are still restricted to rigid-body modelling approaches. At this stage, there is a need for detailed subject-specific models that would allow exploring the links between posture, dynamic stability and muscle function. This study presents the framework and current development of such a model

Efficacy of a fixed combination of permethrin 54.5% and fipronil 6.1% (Effitix®) in dogs experimentally infested with Ixodes ricinus

Abstract Background Ticks are the most important vectors of disease-causing pathogens in domestic animals and are considered to be second worldwide to mosquitoes as vectors of human diseases. In Europe, Ixodes ricinus, the sheep tick, plays an important role as companion animal parasite but is also the primary vector of medically important diseases such as tick-borne encephalitis and Lyme borreliosis. The present study was designed to evaluate the efficacy under laboratory conditions of a new fixed spot-on combination of fipronil and permethrin (Effitix®, Virbac) in treating and preventing tick infestations of Ixodes ricinus in dogs. Methods Twelve dogs were included in this randomized, controlled, blinded laboratory study. They were randomly allocated to two groups of six dogs each according to their pre-treatment live attached Ixodes ricinus tick count. On day 0, the dogs from Group 2 were treated with the recommended dose of Effitix®, the dogs from Group 1 remained untreated. On days −2, 7, 14, 21, 28 and 35, all dogs were infested with 50 (±4) viable unfed adult Ixodes ricinus (20 ± 2 males, 30 ± 2 females). Ticks were removed and counted at 48 ± 2 hours post product administration or tick infestations. Results Through the study, the tick attachment rates for the untreated group were greater than 25% demonstrating that adequate levels of infestation were reached on the control dogs. Based on both arithmetic and geometric means (AM and GM), Effitix® was deemed to be effective against Ixodes ricinus on days 2, 9, 16, 23, 30 and 37 with a percentage of efficacy of 98%, 100%, 100%, 100%, 93% and 95% respectively (AM). No clinical abnormalities were detected during the study. Conclusions The study has shown under laboratory conditions, that Effitix® is a safe and an effective combination to treat and protect dogs from Ixodes ricinus up to 37 days after administration. The high immediate efficacy of 98% evaluated at 48 hours post-treatment was particularly interesting, meaning that Effitix has a curative effect against ticks (Ixodes ricinus) and provides a rapid control of existing Ixodes ricinus infestation on a dog at the time of treatment

Comparaison de critères agronomiques de croissance et de production du cacaoyer en association avec le cocotier

Une étude rationnelle du fonctionnement de l'association cocotier-cacaoyer est entreprise. L'accent est mis sur l'analyse des compétitions, entre les deux plantes, pour l'eau. Les dispositifs étudiés combinent plusieurs densités de plantation des cacaoyers allant du simple )480 plants/ha), double (960 plants/ha) aux triple rangs (1440 plants/ha) dans l'interligne de cocotiers, plantés à densité variable ou fixe (160 plants/ha) selon les expériences. Sur l'une d'elles, de jeunes cacaoyers plantés sur deux rangs sont isolés des cocotiers par une tranchée périphérique de 1 m de profondeur limitant la compétition racinaire. Les critères pris en compte au niveau de la strate cacaoyer sont, pour les cacaoyers adultes, la production et le taux de mortalité, pour les jeunes arbres, la hauteur des troncs, les pourcentages de couronne et la mortalité au champ. Pour de jeunes cacaoyers, les plants sur le traitement " double rangs avec tranchée " sont significativement plus hauts au seuil de 5%, présentent précocement plus de couronnes et ont au taux de remplacement très inférieur à ceux des traitements simple, double et triple rangs. Pour des cacaoyers entrés en production, leur rendement augmente lorsque la densité de plantation des cocotiers diminue. Un suivi régulier de ces critères révèle, dans des conditions hydriques limitante (1994) et non lirnitantes (1995), de fortes compétitions principalement pour J'eau entre plantes associées mais aussi entre cacaoyers selon leurs densités de plantation, et ce, dès le plus jeune âge. Les résultats qui illustrent cette communication sont une étape, complétée en 1996 par la caractérisation du développement aérien et souterrain et l'estimation de la consommation en eau du cacaoyer en association et sur la monoculture. (Résumé d'auteur