Study of pufferfish (Takifugu niphobles) sperm: development of methods for short-term storage, effects of different activation media and role of intracellular changes in Ca2+ and K+ in the initiation of sperm motility

[EN] The first goal of this study was the development of a short-term storage method for pufferfish (Takifugu niphobles) sperm. In this respect, the best results were obtained by diluting the sperm in a seminal-like solution and keeping it in a Petri dish in chilled storage (4 degrees C). This method was successful in preserving sperm quality parameters without resulting in differences in fresh sperm for a relatively long-term period (7 days), for use in aquaculture matters. The addition of bovine serum albumin (BSA) to the medium did not improve the results. On the other hand, both the osmolality and the ion composition of the media are essential factors which can modulate the sperm motility parameters. The osmolality of the activating medium was the most important factor in triggering pufferfish sperm motility, and osmolalities of 750-825 mOsm/kg were necessary to initiate this process, demonstrating that it appears to be a dose-independent mechanism. Regarding the ion composition of the activation media, this study has shown that despite the spermatozoa being able to initiate movement without any ion in the activation medium, the absence of ions can negatively affect the kinetic parameters of the spermatozoa. Finally, in natural conditions (seawater), the activation of sperm motility generates intracellular increases in Ca2+ and K+, suggesting that these ions play an essential role in the activation mechanism of pufferfish sperm. (C) 2013 Elsevier B.V. All rights reserved.This study was funded by the Spanish Ministry of Science and Innovation (MICINN; AGL2010-16009). Victor Gallego has a predoctoral grant (MICINN; BES-2009-020310) and has been granted a fellowship (EEBB-I-12-05858) from the Spanish Personnel Research Training Programme to carry out this study in the Misaki Marine Biological Station (Miura, Japan).Gallego Albiach, V.; Pérez Igualada, LM.; Asturiano Nemesio, JF.; Yoshida, M. (2013). Study of pufferfish (Takifugu niphobles) sperm: development of methods for short-term storage, effects of different activation media and role of intracellular changes in Ca2+ and K+ in the initiation of sperm motility. Aquaculture. 414:82-91. https://doi.org/10.1016/j.aquaculture.2013.07.046S829141

Intracellular changes in Ca2+, K+ and pH after sperm motility activation in the European eel (Anguilla anguilla): Preliminary results

[EN] Although it is widely accepted that osmolality and ion fluxes are the main factors triggering sperm motility in fish, a complex universal mechanism for sperm motility activation does not exist in fish, and studies of marine fish species are even more scarce. Therefore, the main goal of this study was to estimate the intracellular variations in the main ions involved in sperm activation for the first time in European eel, in order to provide additional new data about this little-known process. It was observed that levels of intracellular Ca2+ and K+ sperm ions increased significantly 30 s after the hyperosmotic shock compared to baseline levels, and remained at this level until 120 s post-activation. In contrast, the intracellular pH remained constant during the first 30 s, and decreased gradually at 60 and 120 s post-activation. Our data agree with the current main theory for explaining motility activation in marine fish, in which internal fluctuations of Ca2+ and K+ seem to participate in sperm activation. In addition, fluorescent images showed that both Ca2+ and K+ were concentrated in the apical area of the sperm head, which corresponds to the location of the eel sperm mitochondria, suggesting this organelle plays an important role in sperm motility activation. (C) 2013 Elsevier B.V. All rights reserved.Funded from the European Community's 7th Framework Programme under the Theme 2 "Food, Agriculture and Fisheries, and Biotechnology", grant agreement no 245257 (Pro-Eel) and the Spanish Ministry of Science and Innovation (MICINN; AGL2010-16009). Victor Gallego has a predoctoral grant (MICINN; BES-2009-020310) and has been granted a fellowship (EEBB-I-12-05858) of the Spanish Personnel Research Training Programme to carry out this study in the Universidad de Leon (Leon, Spain). Ilaria Mazzeo had a predoctoral grant from GVA. David S. Penaranda has a contract co-financed by MICINN and UPV (PTA2011-4948-I). F. Martinez-Pastor was supported by the Ramon y Cajal program (RYC-2008-02560, MICINN).Gallego Albiach, V.; Martínez Pastor, F.; Mazzeo, I.; Peñaranda, D.; Herraez, P.; Asturiano Nemesio, JF.; Pérez Igualada, LM. (2014). Intracellular changes in Ca2+, K+ and pH after sperm motility activation in the European eel (Anguilla anguilla): Preliminary results. Aquaculture. 418:155-158. https://doi.org/10.1016/j.aquaculture.2013.10.022S15515841

Sperm motility parameters and spermatozoa morphometric characterization in marine species: a study of swimmer and sessile species

[EN] The biodiversity of marine ecosystems is diverse and a high number of species coexist side by side. However, despite the fact that most of these species share a common fertilization strategy, a high variability in terms of the size, shape, and motion of spermatozoa can be found. In this study, we have analyzed both the sperm motion parameters and the spermatozoa morphometric features of two swimmer (pufferfish and European eel) and two sessile (sea urchin and ascidian) marine species. The most important differences in the sperm motion parameters were registered in the swimming period. Sessile species sperm displayed notably higher values than swimmer species sperm. In addition, the sperm motilities and velocities of the swimmer species decreased sharply once the sperm was activated, whereas the sessile species were able to maintain their initial values for a long time. These results are linked directly to the species-specific lifestyles. Although sessile organisms, which show limited or no movement, need sperm with a capacity to swim for long distances to find the oocytes, swimmer organisms can move toward the female and release gametes near it, and therefore the spermatozoa does not need to swim for such a long time. At the same time, sperm morphology is related to sperm motion parameters, and in this study an in-depth morphometric analysis of ascidian, sea urchin, and pufferfish spermatozoa, using computer-assisted sperm analysis software, has been carried out for the first time. A huge variability in shapes, sizes, and structures of the studied species was found using electron microscopy. (C) 2014 Elsevier Inc. All rights reserved.This study was funded by the Spanish Ministry of Science and Innovation (MICINN; AGL2010-16009). Victor Gallego has a predoctoral grant (MICINN; BES-2009-020310) and has been granted a fellowship (EEBB-I-12-05858) of the Spanish Personnel Research Training Program to carry out this research in the Misaki Marine Biological Station (Miura, Japan).Gallego Albiach, V.; Pérez Igualada, LM.; Asturiano Nemesio, JF.; Yoshida, M. (2014). Sperm motility parameters and spermatozoa morphometric characterization in marine species: a study of swimmer and sessile species. Theriogenology. 82(5):668-676. https://doi.org/10.1016/j.theriogenology.2014.05.026S66867682

Recommandations pour l’utilisation de la toxine botulinique de type A (Botox®) dans l’hyperactivité vésicale réfractaire idiopathique

RésuméObjectifsDéfinir des recommandations pour l’utilisation pratique de la toxine botulinique de type A (BoNTA) dans l’hyperactivité vésicale réfractaire idiopathique (HAVRI).MéthodeÉlaboration de recommandations de bonne pratique par consensus formalisé, validées par un groupe de 13 experts puis par un groupe de lecture indépendant.RésultatsEn cas d’infection urinaire celle-ci doit être traitée et l’injection reportée. Avant l’injection, il est recommandé de s’assurer de la faisabilité et de l’acceptabilité de l’auto-sondage. L’injection peut être réalisée après une anesthésie locale urétro-vésicale (lidocaïne), éventuellement complétée par l’inhalation de protoxyde d’azote et parfois sous anesthésie générale. L’injection sera réalisée au bloc opératoire ou en salle d’endoscopie. La vessie ne doit pas être trop remplie (risque de perforation). Le traitement doit être appliqué en 10 à 20 injections de 0,5 à 1mL réparties de manière homogène dans la vessie en restant à distance des méats urétéraux. Il n’est pas recommandé de laisser en place une sonde vésicale sauf en cas d’hématurie importante. Le patient doit être surveillé jusqu’à la reprise mictionnelle. Une note d’information sur les effets indésirables éventuels doit lui être remise à sa sortie. Une consultation doit être prévue 3 mois après la première injection (calendrier mictionnel, débitmétrie, résidu post-mictionnel et examen cytobactériologique des urines). Un résidu >200mL et/ou symptomatique doit faire discuter des auto-sondages. Une nouvelle injection pourra être envisagée lorsque le bénéfice clinique de la précédente s’estompe (entre 6 et 9 mois).ConclusionsLe respect de ces recommandations devrait permettre une utilisation optimale de la BoNTA.Niveau de preuve3.SummaryObjectivesProvide guidelines for practical usage of botulinum toxin type A (BoNTA) for refractory idiopathic Overactive Bladder management.Patients and methodsGuidelines using formalized consensus guidelines method. These guidelines have been validated by a group of 13 experts quoting proposals, subsequently reviewed by an independent group of experts.ResultsIn the case of patients with urinary tract infection, it must be treated and injection postponed. Before proposing an injection, it is recommended to ensure the feasibility and acceptability of self-catheterisation by patient. The injection can be performed after local anesthesia of the bladder and urethra (lidocaine), supplemented where necessary by nitrous oxide inhalation and sometimes under general anesthesia. Injection is performed in the operating room or endoscopy suite. The bladder should not be too filled (increased risk of perforation). Treatment should be applied in 10 to 20 injections of 0.5 to 1mL homogeneously distributed in the bladder at a distance from the urethral orifices. It is not recommended to leave a urinary catheter in place except in cases of severe hematuria. The patient should be monitored until resumption of micturition. After the first injection, an appointment must be scheduled within 3 months (micturition diary, uroflowmetry, measurement of residual urine and urine culture). Performance of self-catheterisation should be questioned in the case of a symptomatic post-void residual and/or a residue>200mL. A new injection may be considered when the clinical benefit of the previous injection diminishes (between 6 and 9 months). A period of three months must elapse between each injection.ConclusionsImplementation of these guidelines may promote best practice usage of BoNTA with optimal risk/benefit ratio

Comparative genomics of isolates of a pseudomonas aeruginosa epidemic strain associated with chronic lung infections of cystic fibrosis patients

Pseudomonas aeruginosa is the main cause of fatal chronic lung infections among individuals suffering from cystic fibrosis (CF). During the past 15 years, particularly aggressive strains transmitted among CF patients have been identified, initially in Europe and more recently in Canada. The aim of this study was to generate high-quality genome sequences for 7 isolates of the Liverpool epidemic strain (LES) from the United Kingdom and Canada representing different virulence characteristics in order to: (1) associate comparative genomics results with virulence factor variability and (2) identify genomic and/or phenotypic divergence between the two geographical locations. We performed phenotypic characterization of pyoverdine, pyocyanin, motility, biofilm formation, and proteolytic activity. We also assessed the degree of virulence using the Dictyostelium discoideum amoeba model. Comparative genomics analysis revealed at least one large deletion (40-50 kb) in 6 out of the 7 isolates compared to the reference genome of LESB58. These deletions correspond to prophages, which are known to increase the competitiveness of LESB58 in chronic lung infection. We also identified 308 non-synonymous polymorphisms, of which 28 were associated with virulence determinants and 52 with regulatory proteins. At the phenotypic level, isolates showed extensive variability in production of pyocyanin, pyoverdine, proteases and biofilm as well as in swimming motility, while being predominantly avirulent in the amoeba model. Isolates from the two continents were phylogenetically and phenotypically undistinguishable. Most regulatory mutations were isolate-specific and 29% of them were predicted to have high functional impact. Therefore, polymorphism in regulatory genes is likely to be an important basis for phenotypic diversity among LES isolates, which in turn might contribute to this strain's adaptability to varying conditions in the CF lung

Widespread forest vertebrate extinctions induced by a mega hydroelectric dam in lowland Amazonia

Mega hydropower projects in tropical forests pose a major emergent threat to terrestrial and freshwater biodiversity worldwide. Despite the unprecedented number of existing, underconstruction and planned hydroelectric dams in lowland tropical forests, long-term effects on biodiversity have yet to be evaluated. We examine how medium and large-bodied assemblages of terrestrial and arboreal vertebrates (including 35 mammal, bird and tortoise species) responded to the drastic 26-year post-isolation history of archipelagic alteration in landscape structure and habitat quality in a major hydroelectric reservoir of Central Amazonia. The Balbina Hydroelectric Dam inundated 3,129 km2 of primary forests, simultaneously isolating 3,546 land-bridge islands. We conducted intensive biodiversity surveys at 37 of those islands and three adjacent continuous forests using a combination of four survey techniques, and detected strong forest habitat area effects in explaining patterns of vertebrate extinction. Beyond clear area effects, edge-mediated surface fire disturbance was the most important additional driver of species loss, particularly in islands smaller than 10 ha. Based on species-area models, we predict that only 0.7% of all islands now harbor a species-rich vertebrate assemblage consisting of ≥80% of all species. We highlight the colossal erosion in vertebrate diversity driven by a man-made dam and show that the biodiversity impacts of mega dams in lowland tropical forest regions have been severely overlooked. The geopolitical strategy to deploy many more large hydropower infrastructure projects in regions like lowland Amazonia should be urgently reassessed, and we strongly advise that long-term biodiversity impacts should be explicitly included in pre-approval environmental impact assessments