Alteration in neuromuscular function after a 5 km running time trial.

The aim of this study was to characterize the effect of a 5 km running time trial on the neuromuscular properties of the plantar flexors. Eleven well-trained triathletes performed a series of neuromuscular tests before and immediately after the run on a 200 m indoor track. Muscle activation (twitch interpolation) and normalized EMG activity were assessed during maximal voluntary contraction (MVC) of plantar flexors. Maximal soleus H-reflexes and M-waves were evoked at rest (i.e. H (MAX) and M (MAX), respectively) and during MVC (i.e. H (SUP) and M (SUP), respectively). MVC significantly declined (-27%; P < 0.001) after the run, due to decrease in muscle activation (-8%; P < 0.05) and M (MAX)-normalized EMG activity (-13%; P < 0.05). Significant reductions in M-wave amplitudes (M (MAX): -13% and M (SUP): -16%; P < 0.05) as well as H (MAX)/M (MAX) (-37%; P < 0.01) and H (SUP)/M (SUP) (-25%; P < 0.05) ratios occurred with fatigue. Following exercise, the single twitch was characterized by lower peak torque (-16%; P < 0.001) as well as shorter contraction (-19%; P < 0.001) and half-relaxation (-24%; P < 0.001) times. In conclusion, the reduction in plantar flexors strength induced by a 5 km running time trial is caused by peripheral adjustments, which are attributable to a failure of the neuromuscular transmission and excitation-contraction coupling. Fatigue also decreased the magnitude of efferent motor outflow from spinal motor neurons to the plantar flexors and part of this suboptimal neural drive is the result of an inhibition of soleus motoneuron pool reflex excitability.

Almost routine prophylactic cholecystectomy during laparoscopic gastric bypass is safe

Background Morbidly obese patients are at high risk to develop gallstones, and rapid weight loss after bariatric surgery further enhances this risk. The concept of prophylactic cholecystectomy during gastric bypass has been challenged recently because the risk may be lower than reported earlier and because cholecystectomy during laparoscopic gastric bypass may be more difficult and risky. <p>Methods A review of prospectively collected data on 772 patients who underwent laparoscopic primary gastric bypass between January 2000 and August 2007 was performed. The charts of patients operated before 2004 were retrospectively reviewed regarding preoperative echography and histopathological findings.</p> <p>Results Fifty-eight (7.5%) patients had had previous cholecystectomy. In the remaining patients, echography showed gallstones or sludge in 81 (11.3%). Cholecystectomy was performed at the time of gastric bypass in 665 patients (91.7%). Gallstones were found intraoperatively in 25 patients (3.9%), for a total prevalence of gallstones of 21.2%. The age of patients with gallstones was higher than that of gallstone-free patients (43.5 vs 38.7 years, p < 0.0001). Of the removed specimens, 81.8% showed abnormal histologic findings, mainly chronic cholecystitis and cholesterolosis. Cholecystectomy was associated with no procedure-related complication, prolonged duration of surgery by a mean of 19 min (4-45), and had no effect on the duration of hospital stay. Cholecystectomy was deemed too risky in 59 patients (8.3%) who were prescribed a 6-month course of ursodeoxycolic acid.</p> <p>Conclusion Concomitant cholecystectomy can be performed safely in most patients during laparoscopic gastric bypass and does not prolong hospital stay. As such, it is an acceptable form of prophylaxis against stones forming during rapid weight loss. Whether it is superior to chemical prophylaxis remains to be demonstrated in a large prospective randomized study.</p>

Fast analysis of doping agents in urine by ultra-high-pressure liquid chromatography-quadrupole time-of-flight mass spectrometry. II: Confirmatory analysis.

For doping control, analyses of samples are generally achieved in two steps: a rapid screening and, in the case of a positive result, a confirmatory analysis. A two-step methodology based on ultra-high-pressure liquid chromatography coupled to a quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) was developed to screen and confirm 103 doping agents from various classes (e.g., beta-blockers, stimulants, diuretics, and narcotics). The screening method was presented in a previous article as part I (i.e., Fast analysis of doping agents in urine by ultra-high-pressure liquid chromatography-quadrupole time-of-flight mass spectrometry. Part I: screening analysis). For the confirmatory method, basic, neutral and acidic compounds were extracted by a dedicated solid-phase extraction (SPE) in a 96-well plate format and detected by MS in the tandem mode to obtain precursor and characteristic product ions. The mass accuracy and the elemental composition of precursor and product ions were used for compound identification. After validation including matrix effect determination, the method was considered reliable to confirm suspect results without ambiguity according to the positivity criteria established by the World Anti-Doping Agency (WADA). Moreover, an isocratic method was developed to separate ephedrine from its isomer pseudoephedrine and cathine from phenylpropanolamine in a single run, what allowed their direct quantification in urine.

Melt topology and chemistry during partial melting in contact aureoles : an experimental study and field example

This study was initiated to investigate partial melting within the high-grade metamorphic rocks beneath the Little Cottonwood contact aureole (Utah, USA), in order to understand the melt generation, melt migration, and geometry of initial melt distribution on grain scale during crustal anatexis. The emplacement of the Little Cottonwood stock produced a contact aureole in the pelitic host rocks of the Big Cottonwood formation (BC). Metamorphic isogrades in pelitic rocks range form biotite to 2nd sillimanite grade as a function of distance from the contact. Migmatites are restricted to the highest grade and resulted form partial melting of the BC formation rocks. First melt was produced by a combined muscovite/biotite dehydration reaction in the sillimanite + k-feldspar stability field. Melt extraction from the pelites resulted in restites (magnetite + cordierite + alumosilicate ± biotite) surrounded by feldspar enriched quartzite zones. This texture is the result of gradual infiltration of partial melts into the quartzite. Larger, discrete melt accumulation occurred in extensional or transpressional domains such as boudin necks, veins, and ductile shear zones. Melt composition are Si02- rich, crystallized as pegmatites, and apparently were very mobile. They were able to infiltrate the quartzite pervaisivly. These melts are similar in composition to first melts produced in the hydrothermal partial melt experiments at 2kbar between 700 - 800°C on fine grained high metamorphic rocks (andalusite-cordierited-biotite-zone) of the BC formation. The experimental melts are water rich and in disequilibrium with the melting rock. Initial melt composition is heterogeneous for short run duration, reflective a lack of chemical equilibrium between individual melt pools. Rock core scale heterogeneity decreased with time indicating partial homogenization of melt compositions. A simultaneous shift of melt composition to higher silica content with time was observed. The silica content of the melt increased due to local melt/mineral reactions. Melt textures indicate that reactive melt transport is most efficient along grain boundaries rimmed by dissimilar grains. Melt heterogeneity resulted in chemical potential gradients which are major driving forces for initial melt migration and govern melt distribution during initial melting. An additional subject of the thesis is the crystal size distributions of opaque minerals in a fine-grained, high-grade meta-pelite of the Big Cottonwood which were obtained from 3D X-ray tomography (uCT) and 2D thin section analysis. µCT delivers accurate size distributions within a restricted range (~ a factor of 20 in size in a single 3D image), while the absolute number of crystals is difficult to obtain from these sparsely distributed, small crystals on the basis of 2D images. Crystal size distributions obtained from both methods are otherwise similar. - Ce travail de recherche a été entrepris dans le but d'étudier les processus de fusion partielle dans les roches fortement métamorphiques de l'auréole de contact de Little Cottonwood (Utah, USA) et ceci afin de comprendre la génération de liquide de fusion, la migration de ces liquides et la géométrie de la distribution initiale des liquides de fusion à l'échelle du grain durant l'anatexie de la croûte. L'emplacement du petit massif intrusif de Little Cottonwood a produit une auréole de contact dans les roches pélitiques encaissantes appartenant à la Foimation du Big Cottonwood (BC). Les isogrades métamorphiques dans les roches pélitiques varient de l'isograde de la biotite à la deuxième isograde de la sillimanite en fonction de la distance par rapport au massif intrusif. Les migmatites sont restreintes aux zones montrant le plus haut degré métamorphique et résultent de la fusion partielle des roches de la Formation de BC. Le premier liquide de fusion a été produit par la réaction de déshydratation combinée de la muscovite et de la biotite dans le champ de stabilité du feldspath potassique Pt de la sillimanite. L'extraction du liquide de fusion des pélites forme des restites (magnétites + cordiérite + aluminosilicate ± biotite) entourées par des zones de quartzites enrichies en feldspath. Cette texture est le résultat de l'infiltration graduelle du liquide de fusion partielle dans les quartzites. Des accumulations distinctes et plus larges de liquide de fusion ont lieu dans des domaines d'extension ou de transpression tels que les boudins, les veines, et les zones de cisaillement ductile. La composition des liquides de fusion est similaire à celle des liquides pegmatoïdes, et ces liquides sont apparemment très mobiles et capables d'infiltrer les quartzites. Ces liquides de fusion ont la même composition que les premiers liquides produits dans les expériences hydrotheunales de fusion partielle à 2kbar et entre 700-800° C sur les roches finement grenues et hautement métamorphiques (andalousite-cordiérite-biotite zone) de la Formation de BC. Les liquides de fusion obtenus expérimentalement sont riches en eau et sont en déséquilibre avec la roche en fusion. La composition initiale des liquides de fusion est hétérogène pour les expériences de courte durée et reflète l'absence d'équilibre chimique entre les différentes zones d'accumulation des liquides de fusion. L'hétérogénéité à l'échelle du noyau s'estompe avec le temps et témoigne de l'homogénéisation de la composition des liquides de fusion. Par ailleurs, on observe parallèlement un décalage de la composition des liquides vers des compositions plus riches en silice au cours du temps. Le contenu en silice des liquides de fusion évolue vers un liquide pegmatitique en raison des réactions liquides/minéraux. Les textures des liquides de fusion indiquent que le transport des liquides est plus efficace le long des bordures de grains bordés par des grains différents. Aucun changement apparent du volume total n'est visible. L'hétérogénéité des liquides s'accompagne d'un gradient de potentiel chimique qui sert de moteur principal à la migration des liquides et à la distribution des liquides durant la fusion. Un sujet complémentaire de ce travail de thèse réside dans l'étude de la distribution de la taille des cristaux opaques dans les pélites finement grenues et fortement métamorphiques de la Formation de Big Cottonwood. Les distributions de taille ont été obtenues suite à l'analyse d'images 3D acquise par tomographie ainsi que par analyse de lames minces. La microtomographie par rayon X fournit une distribution de taille précise sur une marge restreinte (- un facteur de taille 20 dans une seule image 3D), alors que le nombre absolu de cristaux est difficile à obtenir sur la base d'image 2D en raison de la petite taille et de la faible abondance de ces cristaux. Les distributions de taille obtenues par les deux méthodes sont sinon similaire. Abstact: Chemical differentiation of the primitive Earth was due to melting and separation of melts. Today, melt generation and emplacement is still the dominant process for the growth of the crust. Most granite formation is due to partial melting of the lower crust, followed by transport of magma through the crust to the shallow crust where it is emplaced. Partial melting and melt segregation are essential steps before such a granitic magma can ascent through the crust. The chemistry and physics of partial melting and segregation is complex. Hence detailed studies, in which field observations yield critical information that can be compared to experimental observations, are crucial to the understanding of these fundamental processes that lead and are leading to the chemical stratification of the Earth. The research presented in this thesis is a combined field and experimental study of partial melting of high-grade meta-pelitic rocks of the Little Cottonwood contact aureole (Utah, USA). Contact metamorphic rocks are ideal for textural studies of melt generation, since the relatively short times of the metamorphic event prevents much of the recrystallization which plagues textural studies of lower crustal rocks. The purpose of the study is to characterize melt generation, identify melting reactions, and to constrain melt formation, segregation and migration mechanisms. In parallel an experimental study was undertaken to investigate melt in the high grade meta pelitic rocks, to confirm melt composition, and to compare textures of the partial molten rock cores in the absence of deformation. Results show that a pegmatoidal melt is produced by partial melting of the pelitic rocks. This melt is highly mobile. It is capable of pervasive infiltration of the adjacent quartzite. Infiltration results in rounded quartz grains bordered by a thin feldspar rim. Using computed micro X-ray tomography these melt networks can be imaged. The infiltrated melt leads to rheological weakening and to a decompaction of the solid quartzite. Such decompaction can explain the recent discovery of abundant xenocrysts in many magmas, since it favors the isolation of mineral grains. Pervasive infiltration is apparently strongly influenced by melt viscosity and melt-crystal wetting behavior, both of which depend on the water content of melt and the temperature. In all experiments the first melt is produced on grain boundaries, dominantly by the local minerals. Grain scale heterogeneity of a melting rock leads thus to chemical concentration gradients in the melt, which are the driving force for initial melt migration. Pervasive melt films along grain boundaries leading to an interconnected network are immediately established. The initial chemical heterogeneities in the melt diminish with time. Résumé large public: La différenciation chimique de la Terre primitive est la conséquence de la fusion des roches et de la séparation des liquides qui en résultent. Aujourd'hui, la production de liquide magmatique est toujours le mécanisme dominant pour la croissance de la croûte terrestre. Ainsi la formation de la plupart des granites est un processus qui implique la production de magma par fusion partielle de la croûte inférieure, la migration de ces magmas à travers la croûte et finalement son emplacement dans les niveaux superficielle de la croûte terrestre. Au cours de cette évolution, les processus de fusion partielle et de ségrégation sont des étapes indispensables à l'ascension des granites à travers la croûte. Les conditions physico-chimiques nécessaires à la fusion partielle et à l'extraction de ces liquides sont complexes. C'est pourquoi des études détaillées des processus de fusion partielle sont cruciales pour la compréhension de ces mécanismes fondamentaux responsables de la stratification chimique de la Terre. Parmi ces études, les observations de terrain apportent notamment des informations déterminantes qui peuvent être comparées aux données expérimentales. Le travail de recherche présenté dans ce mémoire de thèse associe études de terrain et données expérimentales sur la fusion partielle des roches pélitiques de haut degré métamorphiques provenant de l'auréole de contact de Little Cottonwood (Utah, USA). Les roches du métamorphisme de contact sont idéales pour l'étude de la folination de liquide de fusion. En effet, la durée relativement courte de ce type d'événement métamorphique prévient en grande partie la recristallisation qui perturbe les études de texture des roches dans la croûte inférieure. Le but de cette étude est de caractériser la génération des liquides de fusion, d'identifier les réactions responsables de la fusion de ces roches et de contraindre la formation de ces liquides et leur mécanisme de ségrégation et de migration. Parallèlement, des travaux expérimentaux ont été entrepris pour reproduire la fusion partielle de ces roches en laboratoire. Cette étude a été effectuée dans le but de confirmer la composition chimique des liquides, et de comparer les textures obtenues en l'absence de déformation. Les résultats montrent qu'un liquide de fusion pegmatoïde est produit par fusion partielle des roches pélitiques. La grande mobilité de ce liquide permet une infiltration pénétrative dans les quarzites. Ces infiltrations se manifestent par des grains de quartz arrondis entourés par une fine bordure de feldspath. L'utilisation de la tomography à rayons X a permis d'obtenir des images de ce réseau de liquide de fusion. L'infiltration de liquide de fusion entraîne un affaiblissement de la rhéologie de la roche ainsi qu'une décompaction des quartzites massifs. Une telle décompaction peut expliquer la découverte récente d'abondants xénocristaux dans beaucoup de magmas, puisque elle favorise l'isolation des minéraux. L'infiltration pénétrative est apparemment fortement influencée par la viscosité du fluide de fusion et le comportement de la tension superficielle entre les cristaux et le liquide, les deux étant dépendant du contenu en eau dans le liquide de fusion et de la température. Dans toutes les expériences, le premier liquide est produit sur les bordures de grains, principalement par les minéraux locaux. L'hétérogénéité à l'échelle des grains d'une roche en fusion conduit donc à un gradient de concentration chimique dans le liquide, qui sert de moteur à l'initiation de la migration du liquide. Des fines couches de liquide de fusion le long de bordures de grains formant un réseau enchevêtré s'établit immédiatement. Les hétérogénéités chimiques initiales dans le liquide s'estompent avec le temps.

Alteration in neuromuscular function after a 5 km running time trial

The aim of this study was to characterize the effect of a 5 km running time trial on the neuromuscular properties of the plantar flexors. Eleven well-trained triathletes performed a series of neuromuscular tests before and immediately after the run on a 200 m indoor track. Muscle activation (twitch interpolation) and normalized EMG activity were assessed during maximal voluntary contraction (MVC) of plantar flexors. Maximal soleus H-reflexes and M-waves were evoked at rest (i.e. H (MAX) and M (MAX), respectively) and during MVC (i.e. H (SUP) and M (SUP), respectively). MVC significantly declined (-27%; P < 0.001) after the run, due to decrease in muscle activation (-8%; P < 0.05) and M (MAX)-normalized EMG activity (-13%; P < 0.05). Significant reductions in M-wave amplitudes (M (MAX): -13% and M (SUP): -16%; P < 0.05) as well as H (MAX)/M (MAX) (-37%; P < 0.01) and H (SUP)/M (SUP) (-25%; P < 0.05) ratios occurred with fatigue. Following exercise, the single twitch was characterized by lower peak torque (-16%; P < 0.001) as well as shorter contraction (-19%; P < 0.001) and half-relaxation (-24%; P < 0.001) times. In conclusion, the reduction in plantar flexors strength induced by a 5 km running time trial is caused by peripheral adjustments, which are attributable to a failure of the neuromuscular transmission and excitation-contraction coupling. Fatigue also decreased the magnitude of efferent motor outflow from spinal motor neurons to the plantar flexors and part of this suboptimal neural drive is the result of an inhibition of soleus motoneuron pool reflex excitability.

Detection of four Plasmodium species in blood from humans by 18S rRNA gene subunit-based and specific real-time PCR assays

Résumé : Un nombre croissant de cas de malaria chez les voyageurs et migrants a été rapporté. Bien que l'analyse microscopique des frottis sanguins reste traditionnellement l'outil diagnostic de référence, sa fiabilité dépend considérablement de l'expertise de l'examinateur, pouvant elle-même faire défaut sous nos latitudes. Une PCR multiplex en temps réel a donc été développée en vue d'une standardisation du diagnostic. Un ensemble d'amorces génériques ciblant une région hautement conservée du gène d'ARN ribosomial 18S du genre Plasmodium a tout d'abord été conçu, dont le polymorphisme du produit d'amplification semblait suffisant pour créer quatre sondes spécifiques à l'espèce P. falciparum, P. malariae, P. vivax et P. ovale. Ces sondes utilisées en PCR en temps réel se sont révélées capables de détecter une seule copie de plasmide de P. falciparum, P. malariae, P. vivax et P. ovale spécifiquement. La même sensibilité a été obtenue avec une sonde de screening pouvant détecter les quatre espèces. Quatre-vingt-dix-sept échantillons de sang ont ensuite été testés, dont on a comparé la microscopie et la PCR en temps réel pour 66 (60 patients) d'entre eux. Ces deux méthodes ont montré une concordance globale de 86% pour la détection de plasmodia. Les résultats discordants ont été réévalués grâce à des données cliniques, une deuxième expertise microscopique et moléculaire (laboratoire de Genève et de l'Institut Suisse Tropical de Bâle), ainsi qu'à l'aide du séquençage. Cette nouvelle analyse s'est prononcé en faveur de la méthode moléculaire pour tous les neuf résultats discordants. Sur les 31 résultats positifs par les deux méthodes, la même réévaluation a pu donner raison 8 fois sur 9 à la PCR en temps réel sur le plan de l'identification de l'espèce plasmodiale. Les 31 autres échantillons ont été analysés pour le suivi de sept patients sous traitement antimalarique. Il a été observé une baisse rapide du nombre de parasites mesurée par la PCR en temps réel chez six des sept patients, baisse correspondant à la parasitémie déterminée microscopiquement. Ceci suggère ainsi le rôle potentiel de la PCR en temps réel dans le suivi thérapeutique des patients traités par antipaludéens. Abstract : There have been reports of increasing numbers of cases of malaria among migrants and travelers. Although microscopic examination of blood smears remains the "gold standard" in diagnosis, this method suffers from insufficient sensitivity and requires considerable expertise. To improve diagnosis, a multiplex real-time PCR was developed. One set of generic primers targeting a highly conserved region of the 18S rRNA gene of the genus Plasmodium was designed; the primer set was polymorphic enough internally to design four species-specific probes for P. falciparum, P. vivax, P. malarie, and P. ovale. Real-time PCR with species-specific probes detected one plasmid copy of P. falciparum, P. vivax, P. malariae, and P. ovale specifically. The same sensitivity was achieved for all species with real-time PCR with the 18S screening probe. Ninety-seven blood samples were investigated. For 66 of them (60 patients), microscopy and real-time PCR results were compared and had a crude agreement of 86% for the detection of plasmodia. Discordant results were reevaluated with clinical, molecular, and sequencing data to resolve them. All nine discordances between 18S screening PCR and microscopy were resolved in favor of the molecular method, as were eight of nine discordances at the species level for the species-specific PCR among the 31 samples positive by both methods. The other 31 blood samples were tested to monitor the antimalaria treatment in seven patients. The number of parasites measured by real-time PCR fell rapidly for six out of seven patients in parallel to parasitemia determined microscopically. This suggests a role of quantitative PCR for the monitoring of patients receiving antimalaria therapy.

Inherently self-calibrating non-Cartesian parallel imaging.

The use of self-calibrating techniques in parallel magnetic resonance imaging eliminates the need for coil sensitivity calibration scans and avoids potential mismatches between calibration scans and subsequent accelerated acquisitions (e.g., as a result of patient motion). Most examples of self-calibrating Cartesian parallel imaging techniques have required the use of modified k-space trajectories that are densely sampled at the center and more sparsely sampled in the periphery. However, spiral and radial trajectories offer inherent self-calibrating characteristics because of their densely sampled center. At no additional cost in acquisition time and with no modification in scanning protocols, in vivo coil sensitivity maps may be extracted from the densely sampled central region of k-space. This work demonstrates the feasibility of self-calibrated spiral and radial parallel imaging using a previously described iterative non-Cartesian sensitivity encoding algorithm.

Quantification of glucuronidated and sulfated steroids in human urine by ultra-high pressure liquid chromatography quadrupole time-of-flight mass spectrometry.

The urinary steroid profile is constituted by anabolic androgenic steroids, including testosterone and its relatives, that are extensively metabolized into phase II sulfated or glucuronidated steroids. The use of liquid chromatography coupled to mass spectrometry (LC-MS) is an issue for the direct analysis of conjugated steroids, which can be used as urinary markers of exogenous steroid administration in doping analysis, without hydrolysis of the conjugated moiety. In this study, a sensitive and selective ultra high-pressure liquid chromatography coupled to quadrupole time-of-flight mass spectrometer (UHPLC-QTOF-MS) method was developed to quantify major urinary metabolites simultaneously after testosterone intake. The sample preparation of the urine (1 mL) was performed by solid-phase extraction on Oasis HLB sorbent using a 96-well plate format. The conjugated steroids were analyzed by UHPLC-QTOF-MS(E) with a single-gradient elution of 36 min (including re-equilibration time) in the negative electrospray ionization mode. MS(E) analysis involved parallel alternating acquisitions of both low- and high-collision energy functions. The method was validated and applied to samples collected from a clinical study performed with a group of healthy human volunteers who had taken testosterone, which were compared with samples from a placebo group. Quantitative results were also compared to GC-MS and LC-MS/MS measurements, and the correlations between data were found appropriate. The acquisition of full mass spectra over the entire mass range with QTOF mass analyzers gives promise of the opportunity to extend the steroid profile to a higher number of conjugated steroids.

Pollution stress on aquatic microbial communities

Thousands of chemical compounds enter the natural environment but many have unknown effects and consequences, in particular at low concentrations. This thesis work contributes to our understanding of pollution effects by using bacteria as test organisms. Bacteria are important for this question because some of them degrade and transform pollutants into less harmful compounds, but secondly because they themselves can be inhibited in their reproduction by exposure to toxic compounds. When inhibitory effects occur this may change the composition of the microbial com¬munity in the long run, leading to altered or diminished ecosystem services by those communities. As a result chemicals of anthropogenic origin may accumulate and per¬sist in the environment, and finally, affect higher organisms as well. In addition to acquiring basic understanding of pollutant effects at low concentrations on bacterial communities an applied goal of this thesis work was to develop bacteria-based tests to screen new organic chemicals for toxicity and biodégradation. In the first part of this work we developed a flow cytometry-based assay on SYT09 plus ethidium-bromide or propidium-iodide stained cells of Pseudomonas ûuorescens exposed or not to a variety of pollutants under oligotrophic growth conditions. Flow cytometry (FC) allows fast and accurate counting of bacterial cells under simul¬taneous assessment of their physiological state, in particular in combination with different fluorescent dyes. Here we employed FC and fluorescent dyes to monitor the effect that pollutants may exert on Pseudomonas ûuorescens SV3. First we designed an oligotrophic growth test, which enabled us to follow population growth at low densities (104 - 10 7 cells per ml) using 0.1 mM sodium acetate as carbon source. Cells in the oligotrophic milieu were then exposed or not to a variety of common pollutants, such as 2-chlorobiphenyl (2CBP), naphthalene (NAH), 4-chlorophenol (4CP), tetradecane (TD), mercury chloride (HgCl2) or benzene, in different dosages. Exposed culture samples were stained with SYT09 (green fluorescent dye binding nucleic acids, generally staining all cells) in combination with propidium iodide (PI) or ethidium bromide (EB), both dyes being membrane integrity indicators. We ob- served that most of the tested compounds decreased population growth in a dosage- dependent manner. SYT09/PI or SYT09/EB staining then revealed that chemical exposure led to arisal of subpopulations of live and injured or dead cells. By modeling population growth on the total cell numbers in population or only the subpopulation of live cells we inferred that even in stressed populations live cells multiply at rates no different to unexposed controls. The net decrease in population growth would thus be a consequence of more and more cells being not able to multiply at all, rather than all cells multiplying at slower rates. In addition, the proportion of injured cells correlated to the compound dosage. We concluded that the oligotrophic test may be useful to asses toxicity of unknown chemicals on a variety of model bacteria. Mul¬tiple tests can be run in parallel and effects are rapidly measured within a period of 8 hours. Interestingly, in the same exposure tests with P. fluorescens SV3 we observed that some chemicals which did not lead to a reduction of net population growth rates did cause measurable effects on live cells. This was mainly observed in cells within the live subpopulation as an increase of the EB fluorescence signal. We showed that SYT09/EB is a more useful combination of dyes than SYT09/PI because PI fluorescence tend to increase only when cells are effectively dead, but not so much in live cells (less then twofold). In contrast, EB geometric mean fluorescence in live cells increased up to eightfold after exposure to toxic compounds. All compounds even at the lowest concentration caused a measurable increase in EB geometric mean fluorescence especially after 2 h incubation time. This effect was found to be transient for cells exposed to 2CBP and 4CP, but chronic for cells incubated with TD and NAH (ultimately leading to cell death). In order to understand the mechanism underlying the observed effects we used known membrane or energy uncouplers. The pattern of EB signal increase in chemical-exposed populations resembled mostly that of EDTA, although EB fluorescence in EDTA-treated or pasteurized cells was even higher than after exposure to the four test chemicals. We conclude that the ability of cells to efflux EB under equilibrium conditions is an appropriate measure for the potential of a chemical to exert toxicity. Since most bacterial species possess efflux systems for EB that all require cellular energy, our test should be more widely relevant to infer toxicity effects of chemical exposure on the physiological status of the bacterial cell. To better understand the effect of toxicant exposure on efflux defense systems, we studied 2-hydroxybiphenyl toxicity to Pseudomonas azeiaica HBP1. We showed that 2-HBP exerts toxicity even to P. azelaica HBP1, but only at concentrations higher than 0.5 mM. Above this concentration transient loss of membrane polarization and integrity occurred, which we conclude from staining of growing cells with fluorescent dyes. Cells finally recover and resume growth on 2HBP. The high resistance of P. azelaica HBP1 to 2-HBP was found to be the result of an efficient MexABOprM- type efflux pump system counteracting passive influx of this compound into the membrane and cellular interior. Mutants with disrupted mexA, mexB and oprM genes did no longer grow on 2-HBP at concentrations above 100 μΜ, whereas below this concentration we found 2-HBP-concentration dependent decrease of growth rate. The MexAB-OprM system in P. azeiaica HBP1 is indeed an efflux pump for ethidium bromide as well. By introducing gfp reporter fusions responsive to intracellular 2- HBP concentrations into HBP1 wild-type or the mutants we demonstrated that 2HBP enters into the cells in a similar way. In contrast, the reporter system in the wild-type cells does not react to 2-HBP at an outside concentration of 2.4 μΜ, whereas in mutant cells it does. This suggests that wild-type cells pump 2-HBP to the outside very effectively preventing accumulation of 2-HBP. 2HBP metabolism, therefore, is not efficient enough to lower the intracellular concentration and prevent toxicity. We conclude that P. azelaica HBP1 resistance to 2-HBP is mainly due to an efficient efflux system and that 2HBP in high concentrations exerts narcotic effects on the bacterial membrane. In the part of this thesis, we investigated the possibilities of bacteria to degrade pollutants at low concentrations (1 mg per L and below). As test components we used 2-hydroxybiphenyl, antibiotics and a variety of fragrances, many of which are known to be difficult to biodegrade. By using accurate counting of low numbers of bacterial cells we could demonstrate that specific growth on these compounds is possible. We demonstrated the accuracy of FC counting at low cell numbers (down to 103 bacterial cells per ml). Then we tested whether bacterial population growth could be specifically monitored at the expense of low substrate concentrations, us¬ing P. azelaica HBP1. A perfect relationship was found between growth rate, yield and 2-HBP concentrations in the range of 0.1 up to 5 mg per L. Mixing P. azelaica within sludge, however, suggested that growth yields in a mixed community can be much lower than in pure culture, perhaps because of loss of metabolic intermediates. We then isolated new strains from activated sludge using 2-HBP or antibiotics (Nal, AMP, SMX) at low concentrations (0.1-1 mg per L) as sole carbon and energy sub¬strate and PAO microdishes. The purified strains were then examined for growth on their respective substrate, which interestingly, showed that all strains can not with¬stand higher than 1 or 10 mg per L concentrations of target substrate. Thus, bacteria must exist that contribute to compound degradation at low pollutant concentrations but are inhibited at higher concentrations. Finally we tested whether specific biomass growth (in number of cells) at the expense of pollutants can also be detected with communities as starting material. Hereto, we focused on a number of fragrance chemicals and measured community biomass increase by flow cytometry cell counting on two distinct starter communities: (i) diluted Lake Geneva water, and dilute activated sludge from a wastewater treatment plant. We observed that most of the test compounds indeed resulted in significant biomass increase in the starter community compared to a no-carbon added control, but activated sludge and lake Geneva water strongly differed (almost mutually ex¬clusive) in their capacity to degrade the test chemicals. In two cases for activated sludge the same type of microbial community developed upon compound exposure, as concluded from transcription fragment length polymorphism analysis on community purified and PCR amplified 16S rRNA gene fragments. To properly test compound biodegradability it is thus important to use starter communities of different origin. We conclude that FC counting can be a valuable tool to screen chemicals for their biodegradability and toxicity. - Des milliers de produits chimiques sont libérés dans l'environnement mais beaucoup ont des effets inconnus, en particulier à basses concentrations. Ce travail de thèse contribue à notre comprehension des effets de la pollution en utilisant des bacteries comme des organismes-tests. Les bacteries sont importantes pour etudier cette ques¬tion car certaines d'entre elles peuvent degrader ou transformer les polluants, mais également parce qu'elles-mmes peuvent tre inhibees dans leur reproduction après avoit ete exposees à ces composes toxiques. Quand des effets inhibiteurs ont lieu, la composition de la communauté microbienne peut tre changee à long terme, ce qui mène à une reduction du service d'ecosystème offert par ces communautés. En consequence, après leur liberation dans l'environnement, les produits chimiques d'origine anthropogenique peuvent soit s'y accumuler et per¬sister, exerant ainsi des effets encore inconnus sur les organismes vivants. En plus d'acquérir des connaissances de base sur les effets des polluants à basses concentra¬tions sur les communautés microbiennes, un but applique de cette thèse était de développer des tests bases sur les bacteries afin d'identifier de nouveau composes pour leur toxicité ou leur biodégradation. Dans la première partie de ce travail, nous avons developpe un test base sur la cytometrie de flux (FC) sur des cellules de Pseudomonas fluorescens colorees par du bromure d'ethidium ou de l'iodure de propidium et exposees ou non à une palette de polluants sous des conditions de croissance oligotrophique. La cytometrie de flux est une technique qui connaît de nombreuses applications dans la microbiologie environ¬nementale. Cela est principalement du au fait qu'elle permet un comptage rapide et precis ainsi que l'évaluation de l'état physiologique, en particulier lorsqu'elle est combinée h des colorations fluorescentes. Ici, nous avons utilise la technique FC et des colorants fluorescents afin de mesurer l'effet que peuvent exercer certains pollu¬ants sur Pseudomonas ûuorescens SV3 . D'abord nous avons conu des tests oligo- trophiques qui nous permettent de suivre la croissance complète de cellules en culture h des densites faibles (104 -10 7 cellules par ml), sur de l'acetate de sodium à 0.1 mM, en presence ou absence de produits chimiques (2-chlorobiphenyl (2CBP), naphthalène (NAH), 4-chlorophenol (4CP), tetradecane (TD), chlorure de mercure(II) (HgCl2)) à différentes concentrations. Afin de montrer le devenir des bacteries tant au niveau de la cellule individuelle que celui de la population globale, après exposition à des series de composes chimiques, nous avons compte les cellules colorees avec du SYT09 (col¬orant fluorescent vert des acides nucléiques pour la discrimination des cellules par rapport au bruit de fond) en combinaison avec l'iodure de propidium (PI) ou le bromure d'ethidium (EB), indicateurs de l'intégrité de la membrane cellulaire avec FC. Nous avons observe que de nombreux composes testes avaient un effet sur la croissance bacterienne, resultant en une baisse du taux de reproduction de la pop¬ulation. En outre, la double coloration que nous avons utilisee dans cette etude SYT09/PI ou SYT09/EB a montre que les produits chimiques testes induisaient une reponse heterogène des cellules dans la population, divisant celle-ci en sous- populations "saine", "endommagee" ou "morte". Les nombres de cellules à partir du comptage et de la proportion de celles "saines" et "endommagees/mortes" ont ensuite ete utilises pour modeliser la croissance de P. ûuorescens SV3 exposee aux produits chimiques. La reduction nette dans la croissance de population est une consequence du fait que de plus en plus de cellules sont incapables de se reproduire, plutt que du fait d'une croissance plus lente de l'ensemble de la population. De plus, la proportion de cellules endommagees est correllee au dosage du compose chimique. Les résultats obtenus nous ont permis de conclure que le test oligotrophique que nous avons developpe peut tre utilise pour l'évaluation de la toxicité de produits chimiques sur différents modèles bacteriens. Des tests multiples peuvent tre lances en parallèle et les effets sont mesures en l'espace de huit heures. Par ailleurs, nous en déduisons que les produits chimiques exercént un effet sur la croissance des cellules de P. ûuorescens SV3, qui est heterogène parmi les cellules dans la population et depend du produit chimique. Il est intéressant de noter que dans les mmes tests d'exposition avec P. ûuorescens SV3, nous avons observe que certains composes qui n'ont pas conduit à une reduction du taux de la croissance nette de la population, ont cause des effets mesurables sur les cellule saines. Ceci a ete essentiellement observe dans la portion "saine" des cellules en tant qu'augmentation du signal de la fluorescence de 1ΈΒ. D'abord nous avons montre que SYT09/EB était une com¬binaison de colorants plus utile que celle de SYT09/PI parce que la fluorescence du PI a tendance à augmenter uniquement lorsque les cellules sont effectivement mortes, et non pas dans les cellules saines (moins de deux fois plus). Par opposi¬tion, la fluorescence moyenne de l'EB dans les cellules saines augmente jusqu'à huit fois plus après exposition aux composes toxiques. Tous les composes, mme aux plus basses concentrations, induisent une augmentation mesurable de la fluorescence moy¬enne de 1ΈΒ, plus particulièrement après deux heures d'incubation. Cet effet s'est revele tre transitoire pour les cellules exposees aux 2CNP et 4CP, mais est chro¬nique pour les cellules incubees avec le TD et le NAH (entranant la mort cellulaire). Afin de comprendre les mécanismes qui sous-tendent les effets observes, nous avons utilise des decoupleurs d'energie ou de membrane. L'augmentation du signal EB dans les populations causee par des produits chimiques ressemblait à celle exerce par le chelateur des ions divalents EDTA. Cependant, les intensités du signal EB des cellules exposees aux produits chimiques testees n'ont jamais atteint les valeurs des cellules traitees avec l'EDTA ou pasteurises. Nous en concluons que le test oli- gotrophique utilisant la coloration (SYT09/)EB des cellules exposees ou non à un produit chimique est utile afin d'evaluer l'effet toxique exerce par les polluants sur la physiologie bacterienne. Afin de mieux comprendre la reaction d'un système de defense par pompe à efflux après exposition à une toxine, nous avons étudié la toxicité du 2-hydroxybiphenyl (2-HBP) sur Pseudomonas azeiaica HBP1. Nous avons montre que le 2-HBP exerce une toxicité mme sur HBP1, mais uniquement à des concentrations supérieures à 0.5 mM. Au-dessus de cette concentration, des pertes transitoires d'intégrité et de polarization membranaire ont lieu, comme cela nous a ete montre par coloration des cellules en croissance. Les cellules sont finalement capables de se rétablir et de reprendre leur croissance sur 2-HBP. La forte resistance de P. azeiaica HBP1 h 2-HBP physiologie bacterienne s'est revele tre le résultat d'un système de pompe h efflux de type MexABOprM qui contre-balance l'influx passif de ce compose h travers la membrane. Nous avons montre, en construisant des mutants avec des insertions dans les gènes mexA, mexB and oprM et des fusions avec le gène rapporteur gfp, que l'altération de n'importe quelle partie du système d'efflux conduisait à accroître l'accumulation de 2-HBP dans la cellule, en comparaison avec la souche sauvage HBP1, provoquant une diminution de la resistance au 2-HBP ainsi qu'une baisse du taux de reproduction des cellules. Des systèmes d'efflux similaires sont répandus chez de nombreuses espèces bactériennes. Ils seraient responsables de la resistance aux produits chimiques tels que les colorants fluorescents (bromure d'ethidium) et des antibiotiques. Nous concluons que la resistance de P. azelaica HBP1 à 2-HBP est principalement due à un système d'efflux efficace et que 2-HBP, à des concentrations elevees, exerce un effet deletère sur la membrane bacterienne. En se basant sur le comptage des cellules avec la FC, nous avons developpe ensuite une methode pour evaluer la biodegradabilite de polluants tels que le 2-HBP ainsi que les antibiotiques (acide nalidixique (Nal), ampicilline (AMP) ou sulfamethoxazole (SMX)) à de faibles concentrations lmg par L et moins), par le suivi de la croissance spécifique sur le compose de cultures microbiennes pures et mixtes. En utilisant un comptage precis de faibles quantités de cellules nous avons pu demontrer que la croissance spécifique sur ces composes est possible. Nous avons pu illustrer la precision du comptage par cytometrie de flux à faible quantité de cellules (jusqu'à 10 3 cellules par ml). Ensuite, nous avons teste s'il était possible de suivre dynamiquement la croissance de la population de cellules sur faibles concentrations de substrats, en utilisant P. azelaica HBP1. Une relation parfaite a ete trouvee entre le taux de croissance, le rendement et les concentrations de 2-HBP (entre 0.1 et 5 mg par L). En mélangeant HBP1 à de la boue active, nous avons pu montrer que le rendement en communauté mixtes pouvait tre bien inférieur qu'en culture pure. Ceci étant peut tre le résultat d'une perte d'intermédiaires métaboliques. Nous avons ensuite isole de nouvelles souches à partir de la boue active en utilisant le 2-HBP ou des antibiotiques (Nal, AMP, SMX) h basses concentrations (0.1-1 mg par L) comme seules sources de carbone et d'energie. En combinaison avec ceci, nous avons également utilise des microplaques PAO. Les souches purifiees ont ensuite ete examinees pour leurs croissances sur leurs substrats respectifs. De faon intéressante, toutes ces souches ont montre qu'elles ne pouvaient pas survivre à des concentrations de substrats supérieures à 1 ou 10 mg par L. Ainsi, il existe des bacteries qui contribuent à la degradation de composes à basses concentrations de polluant mais sont inhibes lorsque ces concentrations deviennent plus hautes. Finalement, nous avons cherche à savoir s'il est possible de detecter une croissance spécifique à une biomasse au depend d'un polluant, en partant d'une communauté microbienne. Ainsi, nous nous sommes concentre sur certains composes et avons mesure l'augmentation de la biomasse d'une communauté grce à la cytometrie de flux. Nous avons compte deux communautés de depart distinctes: (i) une dilution d'eau du Lac Léman, et une dilution de boue active d'une station d'épuration. Nous avons observe que la plupart des composes testes ont entrane une augmentation de la biomasse de depart par rapport au control sans addition de source de carbone. Néanmoins, les échantillons du lac Léman et de la station d'épuration différaient largement (s'excluant mutuellement l'un l'autre) dans leur capacité à degrader les composes chimiques. Dans deux cas provenant de la station d'épuration, le mme type de communauté microbienne s'est developpe après exposition aux composes, comme l'a démontré l'analyse TRFLP sur les fragments d'ARN 16S purifie de la communauté et amplifie par PCR. Afin de tester correctement la biodegradabilite d'un compose, il est donc important d'utiliser des communautés de depart de différentes origines Nous en concluons que le comptage par cytometrie de flux peut tre un outil de grande utilité pour mettre en valeur la biodegradabillite et la toxicité des composes chimiques.

Changes in leg-spring behavior during a 5000 m self-paced run in differently trained athletes

Summary Aims.-To explore whether fatigue-induced changes in spring-mass behavior during a 5000m self-paced run varied according to the runner's training status. Methods and results.-Six highly- and six well-trained triathletes completed a 5000m time trial. Running velocity and vertical stiffness decreased significantly (P < 0.05) with fatigue, whereas leg stiffness remained constant. None of these parameters displayed a significant interaction between fatigue and training status, despite vertical stiffness being higher (P < 0.05) in highly-trained triathletes. Conclusions.-During a 5000m self-paced run, impairments in leg-spring behavior that occur with fatigue are not affected by athletes' training status. © 2009 Elsevier Masson SAS. All rights reserved. Objectifs.-Étudier, chez des athlètes de niveaux différents, les modifications de raideur mécanique liées à l'apparition de la fatigue lors d'une course de 5000 m. Synthèse des faits.-Six triathlètes très entraînés et six autres bien entraînés ont réalisé une course de 5000 m. La vitesse de course et la raideur verticale diminuaient significativement (p < 0,05) avec la fatigue, alors que la raideur de la jambe demeurait inchangée. Aucune interaction entre la fatigue et le niveau d'entraînement n'a été détectée, malgré des niveaux de raideur verticale plus élevés (p < 0,05) chez les sujets les mieux entraînés.

Changes in leg spring behaviour, plantar loading and foot mobility magnitude induced by an exhaustive treadmill run in adolescent middle-distance runners.

OBJECTIVES: This study aimed to determine adjustments in spring-mass model characteristics, plantar loading and foot mobility induced by an exhaustive run. DESIGN: Within-participants repeated measures. METHODS: Eleven highly-trained adolescent middle-distance runners ran to exhaustion on a treadmill at a constant velocity corresponding to 95% of velocity associated with VO₂max (17.8 ± 1.4 kmh(-1), time to exhaustion=8.8 ± 3.4 min). Contact time obtained from plantar pressure sensors was used to estimate spring-mass model characteristics, which were recorded (during 30 s) 1 min after the start and prior to exhaustion using pressure insoles. Foot mobility magnitude (a composite measure of vertical and medial-lateral mobility of the midfoot) was measured before and after the run. RESULTS: Mean contact area (foot to ground), contact time, peak vertical ground reaction force, centre of mass vertical displacement and leg compression increased significantly with fatigue, while flight time, leg stiffness and mean pressure decreased. Leg stiffness decreased because leg compression increased to a larger extent than peak vertical ground reaction forces. Step length, step frequency and foot mobility magnitude did not change at exhaustion. CONCLUSIONS: The stride pattern of adolescents when running on a treadmill at high constant velocity deteriorates near exhaustion, as evidenced by impaired leg-spring behaviour (leg stiffness) and altered plantar loading.

Is ultracision knife safe and efficient for breast capsulectomy? A preliminary study.

BACKGROUND: Silicone breast implants are used to a wide extent in the field of plastic surgery. However, capsular contracture remains a considerable concern. This study aimed to analyze the effectiveness and applicability of an ultracision knife for capsulectomy breast surgery. METHODS: A prospective, single-center, randomized study was performed in 2009. The inclusion criteria specified female patients 20-80 years of age with capsular contracture (Baker 3-4). Ventral capsulectomy was performed using an ultracision knife on one side and the conventional Metzenbaum-type scissors and surgical knife on the collateral side of the breast. Measurements of the resected capsular ventral fragment, operative time, remaining breast tissue, drainage time, seroma and hematoma formation, visual analog scale pain score, and sensory function of the nipple-areola complex were assessed. In addition, histologic analysis of the resected capsule was performed. RESULTS: Five patients (median age, 59.2 years) were included in this study with a mean follow-up period of 6 months. Three patients had Baker grade 3 capsular contracture, and two patients had Baker grade 4 capsular contracture. The ultracision knife was associated with a significantly lower pain score, shorter operative time, smaller drainage volume, and shorter drainage time and resulted in a larger amount of remaining breast tissue. Histologic analysis of the resected capsule showed no apoptotic cells in the study group or control group. CONCLUSIONS: The results suggest that ventral capsulectomy with Baker grade 3 or 4 contracture using the ultracision knife is feasible, safe, and more efficient than blunt dissection and monopolar cutting diathermy and has a short learning curve. LEVEL OF EVIDENCE II: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors at www.springer.com/00266 .

Low-dose valganciclovir is efficacious and safe for prophylaxis of cytomegalovirus infection in kidney transplantation

Background: Optimal valganciclovir (VGC) dosage and duration for cytomegalovirus (CMV) prophylaxis in kidney transplant recipients remains controversial. This study aimed to determine GCV blood levels and efficacy/safety observed under low-dose oral VGC in kidney transplant recipients. Secondly, to quantify the variability of GCV blood levels, and its potential clinical impact. Methods: In this prospective study, each patient at risk for CMV undergoing kidney transplantation received low-dose VGC (450 mg qd) prophylaxis for 3 months, unless GFR was below 40 mL/min, in which case the dose was adapted to 450 mg every other day. GCV levels, at trough (Ctrough) and at peak (C3h) were measured monthly and CMV viremia was assessed during and after prophylaxis using real time quantitative Polymerase Chain Reaction. Adverse effects were recorded on each GCV sampling. Patients were followed up to one year after transplantation. Results: 38 kidney recipients (19 D+/R+, 11 D+/R-, 8 D-/R+) received 3-month VGC prophylaxis. Most patients (mean GFR of 59 mL/min) received 450 mg qd but the dose was reduced to 450 mg every other day in 6 patients with mean GFR of 22 mL/min. Average GCV C3h and Ctrough (regressed at 24h or 48h) were 3.9 mg/L (CV 33%, range: 1.3-8.2) and 0.4 mg/L (CV 111%, range 0.1-3.3). Population pharmacokinetic analysis showed a fair dispersion of the parameters mainly influenced by renal function. Despite this variability, patients remained aviremic during VGC prophylaxis. Neutropenia and thrombocytopenia (grade 2-4) were reported in 4% and 3% of patients respectively. During follow-up, asymptomatic CMV viremia was reported in 25% patients. One year after transplantation, 12% patients (all D+/R-) had developed a CMV disease, which was treated with a therapeutic 6-week course of oral VGC. Conclusion: Average GCV blood levels after oral administration of low-dose VGC in kidney transplant recipients were comparable to those previously reported with oral GCV prophylaxis, efficacious and well tolerated. Thus, a 3-month course of low-dose VGC is appropriate for the renal function of most kidney transplant recipients.

Development of a real-time PCR for the specific detection of Waddlia chondrophila in clinical samples.

Waddlia chondrophila is considered as an emerging human pathogen likely involved in miscarriage and lower respiratory tract infections. Given the low sensitivity of cell culture to recover such an obligate intracellular bacteria, molecular-based diagnostic approaches are warranted. We thus developed a real-time PCR that amplifies Waddlia chondrophila DNA. Specific primers and probe were selected to target the 16S rRNA gene. The PCR specifically amplified W. chondrophila but did not amplify other related-bacteria such as Parachlamydia acanthamoebae, Simkania negevensis and Chlamydia pneumoniae. The PCR exhibited a good intra-run and inter-run reproducibility and a sensitivity of less than ten copies of the positive control. This real-time PCR was then applied to 32 nasopharyngeal aspirates taken from children with bronchiolitis not due to respiratory syncytial virus (RSV). Three samples revealed to be Waddlia positive, suggesting a possible role of this Chlamydia-related bacteria in this setting.

Aerosol immunization with NYVAC and MVA vectored vaccines is safe, simple, and immunogenic.

Each year, approximately five million people die worldwide from putatively vaccine-preventable mucosally transmitted diseases. With respect to mass vaccination campaigns, one strategy to cope with this formidable challenge is aerosol vaccine delivery, which offers potential safety, logistical, and cost-saving advantages over traditional vaccination routes. Additionally, aerosol vaccination may elicit pivotal mucosal immune responses that could contain or eliminate mucosally transmitted pathogens in a preventative or therapeutic vaccine context. In this current preclinical non-human primate investigation, we demonstrate the feasibility of aerosol vaccination with the recombinant poxvirus-based vaccine vectors NYVAC and MVA. Real-time in vivo scintigraphy experiments with radiolabeled, aerosol-administered NYVAC-C (Clade C, HIV-1 vaccine) and MVA-HPV vaccines revealed consistent mucosal delivery to the respiratory tract. Furthermore, aerosol delivery of the vaccines was safe, inducing no vaccine-associated pathology, in particular in the brain and lungs, and was immunogenic. Administration of a DNA-C/NYVAC-C prime/boost regime resulted in both systemic and anal-genital HIV-specific immune responses that were still detectable 5 months after immunization. Thus, aerosol vaccination with NYVAC and MVA vectored vaccines constitutes a tool for large-scale vaccine efforts against mucosally transmitted pathogens.