Impact of Treatment with Unilateral Implant- Supported Fixed Partial Prosthesis on the Frequency of Masticatory Side Switches in Patients with Unilateral Posterior Missing Teeth.

This study aimed to determine the impact of treatment with an implant-supported fixed partial prosthesis (ISFPP) on the frequency of masticatory side switches in patients with unilateral posterior missing teeth. This was a prospective study of 30 patients with unilateral posterior missing teeth treated with one-, two-, or three-unit ISFPPs. Comparison was with 10 healthy individuals with complete natural dentitions. Each participant performed masticatory assays, which involved chewing pieces of silicon inside a latex bag, at baseline and at 3-months' follow-up. The frequency of masticatory side switches was reported as the masticatory side-switch index: the number of side switches divided by the maximum number of possible switches. Data were analyzed by Kruskal-Wallis, Mann-Whitney U test, or Wilcoxon test, as appropriate. At baseline, the masticatory side-switch index was lower only in patients with three missing teeth than in controls. At 3 months after treatment, a significant increase in the masticatory side-switch index was only observed in patients treated with three restorative units. Treatment with ISFPPs does not change the masticatory side-switch frequency in patients with unilateral posterior missing teeth, though it could increase it in patients with three missing teeth.

The addition of Lactobacillus spp. negatively affects Mycoplasma bovis viability in bovine cervical mucus.

BACKGROUND: Mycoplasma bovis is an important pathogen for the cattle industry worldwide causing significant economic losses. Several transmission routes, including those related to reproduction, have been described. Indeed, the pathogen can colonize the female reproductive tract after artificial insemination (AI) with contaminated semen. Lactobacillus spp.-based probiotics have been used for vaginal dysbiosis treatment in women and cows although their role in controlling cervico-vaginal infections due to M. bovis is unknown. The objective of the present work is to assess the viability of M. bovis (PG45, NCTC 10131) in experimentally contaminated cervical mucus after the addition of Lactobacillus spp. at different concentrations as a competing agent and pH acidifier. RESULTS: The addition of probiotic at a concentration higher than 108 colony forming units (CFU/mL had a detrimental effect (P < 0.05) on mycoplasma viability in cervical mucus. This coincided with a significant LAB growth and an important decrease in pH from 8.4 to 5.6 (P < 0.05). However, after the addition of less concentrated probiotic, M. bovis survival was not affected and there was no significant LAB growth despite the drop of pH from 8.4 to 6.73 (P < 0.05). CONCLUSION: The addition of concentrations higher than 108 CFU/mL of Lactobacillus spp. negatively affects M. bovis viability in bovine cervical mucus under in vitro conditions. Although the effect observed on the pathogen viability seems to be related to the pH decrease after LAB proliferation in cervical mucus, further studies are necessary to elucidate if other factors are implicated. Nevertheless, the administration of Lactobacillus spp.-based probiotics might be used in the future to control M. bovis proliferation in the cervico-vaginal tract of cows.

Good farming practices in apiculture.

Modern European beekeeping is facing numerous challenges due to a variety of factors, mainly related to globalisation, agrochemical pollution and environmental changes. In addition to this, new pathogens threaten the health of European honey bees. In that context, correct colony management should encompass a wider vision, where productivity aspects are linked to a One Health approach in order to protect honey bees, humans and the environment. This paper describes a novel tool to be applied in beekeeping operations: good beekeeping practices (GBPs). The authors ranked a list of GBPs scored against their importance and validated by an international team, including researchers, national animal health authorities and international beekeepers' associations. These activities were carried out in the project 'BPRACTICES', approved within the transnational call of the European Research Area Network on Sustainable Animal Production (ERA-NET SusAn) in the Horizon 2020 Research and Innovation Programme of the European Union. This study, created through an international collaboration, aims to present an innovative and implementable approach, similar to applications already adopted in other livestock production systems.

L'apiculture moderne européenne est confrontée à de nombreuses difficultés dues à divers facteurs, pour la plupart liés à la mondialisation, à la pollution agrochimique et à la modification de l'environnement. À ces facteurs s'ajoute l'émergence de nouveaux agents pathogènes qui menacent la santé des abeilles mellifères d'Europe. Dans ce contexte, une gestion appropriée des colonies d'abeilles devrait reposer sur une vision plus large, dans laquelle les aspects relevant de la productivité sont examinés suivant une approche « Une seule santé ¼ afin de protéger les abeilles mellifères, les humains et l'environnement. Les auteurs décrivent un nouvel outil destiné à l'apiculture : les bonnes pratiques apicoles. Ils ont évalué et classé par ordre d'importance une liste de bonnes pratiques apicoles validées par une équipe internationale composée de chercheurs, d'autorités nationales de la santé animale et d'associations internationales d'apiculteurs. Ces activités ont été conduites dans le cadre du projet « BPRACTICES ¼, proposition retenue suite à l'appel à projets transnationaux du réseau ERA­NET SusAn (European Research Area Network on Sustainable Animal Production) au sein du Programme Horizon 2020 de l'Union européenne pour la recherche et l'innovation. Conçue sous forme de collaboration internationale, cette étude vise à proposer une approche innovante et pratique, similaire aux applications précédemment adoptées dans d'autres systèmes de production animale.

La apicultura europea hace frente a numerosos problemas resultantes de diversos factores, relacionados principalmente con la mundialización, la contaminación agroquímica y los cambios ambientales, a todo lo cual se suman nuevos patógenos que amenazan la salud de las abejas melíferas europeas. En este contexto, una correcta gestión de las colonias debe traer aparejada una visión más global, en la que las cuestiones de productividad se consideren en clave de «Una sola salud¼ con objeto de proteger tanto a las abejas melíferas como a las personas y el medio ambiente. En este artículo se describe una novedosa herramienta aplicable a la actividad apícola: las buenas prácticas de apicultura. Los autores jerarquizaron una serie de buenas prácticas de apicultura seleccionadas, validadas y puntuadas según su importancia por un equipo internacional que incluía a investigadores, autoridades nacionales de sanidad animal y asociaciones internacionales de apicultores. Este trabajo formaba parte del proyecto «BPRACTICES¼, aprobado con ocasión de la convocatoria internacional abierta por la Red del espacio europeo de investigación en sanidad animal sostenible (ERA­NET SusAn), inscrita a su vez en Horizonte 2020, el programa de investigación e innovación de la Unión Europea. El estudio aquí descrito, fruto de la colaboración internacional, tiene por objeto presentar un planteamiento novedoso y viable, parecido a las aplicaciones ya implantadas en otros sistemas de producción animal.

Direct detection of the triphenylpyrylium-derived short-lived intermediates in the photocatalyzed degradation of acetaminophen, acetamiprid, caffeine and carbamazepine.

Advanced oxidation processes are useful methodologies to accomplish abatement of contaminants; however, elucidation of the reaction mechanisms is hampered by the difficult detection of the short-lived primary key species involved in the photocatalytic processes. Nevertheless, herein the combined use of an organic photocatalyst such as triphenylpyrylium (TPP+) and photophysical techniques based on emission and absorption spectroscopy allowed monitoring the photocatalyst-derived short-lived intermediates. This methodology has been applied to the photocatalyzed degradation of different pollutants, such as acetaminophen, acetamiprid, caffeine and carbamazepine. First, photocatalytic degradation of a mixture of the pollutants showed that acetaminophen was the most easily photodegraded, followed by carbamazepine and caffeine, being the abatement of acetamiprid almost negligible. This process was accompanied by mineralization, as demonstrated by trapping of carbon dioxide using barium hydroxide. Then, emission spectroscopy measurements (steady-state and time-resolved fluorescence) allowed demonstrating quenching of the singlet excited state of TPP+. Laser flash photolysis experiments with absorption detection showed that oxidation of contaminants is accompanied by TPP+ reduction, with formation of a pyranyl radical (TPP), that constituted a fingerprint of the redox nature of the occurring process. The relative amounts of TPP detected was also correlated with the efficiency of the photodegradation process.

Direct detection of the triphenylpyrylium-derived short-lived intermediates in the photocatalyzed degradation of acetaminophen, acetamiprid, caffeine and carbamazepine.

Advanced oxidation processes are useful methodologies to accomplish abatement of contaminants; however, elucidation of the reaction mechanisms is hampered by the difficult detection of the short-lived primary key species involved in the photocatalytic processes. Nevertheless, herein the combined use of an organic photocatalyst such as triphenylpyrylium (TPP+) and photophysical techniques based on emission and absorption spectroscopy allowed monitoring the photocatalyst-derived short-lived intermediates. This methodology has been applied to the photocatalyzed degradation of different pollutants, such as acetaminophen, acetamiprid, caffeine and carbamazepine. First, photocatalytic degradation of a mixture of the pollutants showed that acetaminophen was the most easily photodegraded, followed by carbamazepine and caffeine, being the abatement of acetamiprid almost negligible. This process was accompanied by mineralization, as demonstrated by trapping of carbon dioxide using barium hydroxide. Then, emission spectroscopy measurements (steady-state and time-resolved fluorescence) allowed demonstrating quenching of the singlet excited state of TPP+. Laser flash photolysis experiments with absorption detection showed that oxidation of contaminants is accompanied by TPP+ reduction, with formation of a pyranyl radical (TPP), that constituted a fingerprint of the redox nature of the occurring process. The relative amounts of TPP detected was also correlated with the efficiency of the photodegradation process.

Functional limitation of the masticatory system in patients with bulbar involvement in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) with bulbar dysfunction affects the motor neurons responsible for controlling the muscles in the jaw, face, soft palate, pharynx, larynx and tongue. This cross-sectional study aimed to determine the functional limitation of the jaw in patients with ALS and bulbar dysfunction who had upper motor neuron (UMN), lower motor neuron (LMN) or balanced involvement. One hundred and fifty-three patients with ALS and 23 controls were included. All participants answered using the 8-item Jaw Functional Limitation Scale (JFLS-8). Patients with ALS were grouped by neurologic examination as follows: non-bulbar ALS, bulbar UMN-predominant ALS; bulbar LMN-predominant ALS; and bulbar balanced (UMN + LMN) ALS. Jaw limitation between the different groups was compared using the Kruskal-Wallis test. Patients with non-bulbar ALS had similar mandibular limitations to healthy participants. Only patients with balanced UMN and LMN bulbar manifestations reported greater difficulties in chewing soft food or in jaw mobility compared to the non-bulbar ALS group. Patients with bulbar involvement also had greater difficulties in chewing tough food or chicken and in swallowing and talking compared to the non-bulbar group, regardless of whether UMN or LMN predominant. No significant differences were found between the groups in smiling and yawning difficulties. Bulbar involvement in patients with ALS is associated with functional limitation of the masticatory system. However, balanced bulbar UMN and LMN involvement is associated with the worst impairments in chewing soft food and in opening the jaw widely.

Correction: Safety of Seasonal Malaria Chemoprevention (SMC) with Sulfadoxine-Pyrimethamine plus Amodiaquine when Delivered to Children under 10 Years of Age by District Health Services in Senegal: Results from a Stepped-Wedge Cluster Randomized Trial.

[This corrects the article DOI: 10.1371/journal.pone.0162563.].

Safety of Seasonal Malaria Chemoprevention (SMC) with Sulfadoxine-Pyrimethamine plus Amodiaquine when Delivered to Children under 10 Years of Age by District Health Services in Senegal: Results from a Stepped-Wedge Cluster Randomized Trial.

BACKGROUND: It is recommended that children aged 3 months to five years of age living in areas of seasonal transmission in the sub-Sahel should receive Seasonal Malaria Chemoprevention (SMC) with sulfadoxine-pyrimethamine plus amodiaquine (SPAQ) during the malaria transmission season. The purpose of this study was to evaluate the safety of SMC with SPAQ in children when delivered by community health workers in three districts in Senegal where SMC was introduced over three years, in children from 3 months of age to five years of age in the first year, then in children up to 10 years of age. METHODS: A surveillance system was established to record all deaths and all malaria cases diagnosed at health facilities and a pharmacovigilance system was established to detect adverse drug reactions. Health posts were randomized to introduce SMC in a stepped wedge design. SMC with SPAQ was administered once per month from September to November, by nine health-posts in 2008, by 27 in 2009 and by 45 in 2010. RESULTS: After three years, 780,000 documented courses of SMC had been administered. High coverage was achieved. No serious adverse events attributable to the intervention were detected, despite a high level of surveillance. CONCLUSIONS: SMC is being implemented in countries of the sub-Sahel for children under 5 years of age, but in some areas the age distribution of cases of malaria may justify extending this age limit, as has been done in Senegal. Our results show that SMC is well tolerated in children under five and in older children. However, pharmacovigilance should be maintained where SMC is implemented and provision for strengthening national pharmacovigilance systems should be included in plans for SMC implementation. TRIAL REGISTRATION: ClinicalTrials.gov NCT 00712374.

[Identification of anopheles breeding sites in the residual foci of low malaria transmission «hotspots¼ in Central and Western Senegal]. / Identification des gîtes larvaires d'anophèles dans les foyers résiduels de faible transmission du paludisme « hotspots ¼ au centre-ouest du Sénégal.

Malaria incidence has markedly declined in the Mbour, Fatick, Niakhar and Bambey districts (central and western Senegal) thanks to a scaling up of effective control measures namely LLINs (Long Lasting Insecticide Treated Net), ACTs (Artesunate Combination Therapy) and promoting care seeking. However malaria cases are now maintained by foci of transmission called hotspots. We evaluate the role of anopheles breeding sites in the identification of malaria hotspots in the health districts of Mbour, Fatick, Niakhar and Bambey. Surveys of breeding sites were made in 6 hotspot villages and 4 non-hotspot villages. A sample was taken in each water point with mosquito larvae by dipping method and the collected specimens were identified to the genus level. Additional parameters as name of the village and breeding sites, type of collection, original water turbidity, presence of vegetation, proximity to dwellings, geographic coordinates, sizes were also collected. Sixty-two water collections were surveyed and monitored between 2013 and 2014. Temporary natural breeding sites were predominant regardless of the epidemiological status of the village. Among the 31 breeding sites located within 500 meters of dwellings in hotspots villages, 70% carried Anopheles larvae during the rainy season while 43% of the 21 breeding sites located at similar distances in non-hotspot villages carried Anopheles larvae during the same period (P = 0.042). At the end of the rainy season, the trend is the same with 27% of positive breeding sites in hotspots and 14% in non-hotspots villages. The breeding sites encountered in hotspots villages are mostly small to medium size and are more productive by Anopheles larvae than those found in non-hotspot area. This study showed that the high frequency of smallest and productive breeding sites around and inside the villages can create conditions of residual transmission.

Peripheral, functional and postural asymmetries related to the preferred chewing side in adults with natural dentition.

The aim of this cross-sectional study was to determine the preferred chewing side and whether chewing side preference is related to peripheral, functional or postural lateral preferences. One hundred and forty-six adults with natural dentition performed three masticatory assays, each consisting of five trials of chewing three pieces of silicon placed into a latex bag for 20 cycles, either freestyle or unilaterally on the right- or left-hand side. Occlusal contact area in the intercuspal position, maximum bite force, masticatory performance and cycle duration were measured and the lateral asymmetry of these variables was calculated. Laterality tests were performed to determine handedness, footedness, earedness and eyedness as functional preferences, and hand-clasping, arm-folding and leg-crossing as postural lateral preferences. The preferred chewing side was determined using three different methods: assessment of the first chewing cycle for each trial, calculation of the asymmetry index from all cycles and application of a visual analogue scale. Bivariate relationship and multiple linear regression analyses were performed. Among unilateral chewers, 77% of them preferred the right side for chewing. The factors most closely related to the preferred chewing side were asymmetry of bite force, asymmetry of masticatory performance and earedness, which explained up to 16% of the variance. Although several functional or postural lateral preferences seem to be related to the preferred chewing side, peripheral factors such as asymmetry of bite force and of masticatory performance are the most closely related to the preferred chewing side in adults with natural dentition.

[Acceptability of indoor residual spraying in the Central-Western of Senegal]. / Étude de l'acceptabilité de l'aspersion intradomiciliaire d'insecticide à effet rémanent dans le centre-ouest du Sénégal.

The recent decline of malaria transmission in central-western of Senegal after a scaling up of control measures gives an open window for interventions toward malaria elimination. As a consequence, malaria transmission is now occurring as hotspots. The aim of the project is to evaluate whether target control measures combining indoor residual spraying (IRS) with chemoprophylaxis can virtually eliminate malaria in hotspots. Targeted villages located in four (4) health districts (Mbour, Fatick, Niakhar and Bambey) were sprayed in august 2013 with Actellic® 300 CS (Pirimiphosmethyl). Our objective in this study is to evaluate the acceptability of IRS in the population. IRS is a very complex intervention that requires strong adhesion of populations. After its implementation, 370 households have been interviewed. The results of this survey showed good acceptability of IRS using Actellic® 300 CS, with 97.8% of beneficiaries who declared that IRS is good and even excellent for the community. Despite inconveniences that may arise during intervention, including the preparation of structures to be treated, 98% of respondents were not disturbed in their daily activities. 98.6% of responders declared that sprayers were working with professionalism and almost all households (99.7%) are willing to accept IRS next year. The survey revealed a good acceptability of indoor residual spraying in hot spots located in central-western of Senegal; spraying with Actellic® 300 CS did not cause a problem to the targeted populations. Finally, there is great satisfaction in the population due a huge reduction of mosquito nuisances.

The effects of superovulation of donor sows on ovarian response and embryo development after nonsurgical deep-uterine embryo transfer.

This study aimed to evaluate the effectiveness of superovulation protocols in improving the efficiency of embryo donors for porcine nonsurgical deep-uterine (NsDU) embryo transfer (ET) programs. After weaning (24 hours), purebred Duroc sows (2-6 parity) were treated with 1000 IU (n = 27) or 1500 IU (n = 27) of eCG. Only sows with clear signs of estrus 4 to 72 hours after eCG administration were treated with 750 IU hCG at the onset of estrus. Nonhormonally treated postweaning estrus sows (n = 36) were used as a control. Sows were inseminated and subjected to laparotomy on Days 5 to 6 (Day 0 = onset of estrus). Three sows (11.1%) treated with the highest dosage of eCG presented with polycystic ovaries without signs of ovulation. The remaining sows from nonsuperovulated and superovulated groups were all pregnant, with no differences in fertilization rates among groups. The number of CLs and viable embryos was higher (P < 0.05) in the superovulated groups compared with the controls and increased (P < 0.05) with increasing doses of eCG. There were no differences among groups in the number of oocytes and/or degenerated embryos. The number of transferable embryos (morulae and unhatched blastocysts) obtained in pregnant sows was higher (P < 0.05) in the superovulated groups than in the control group. In all groups, there was a significant correlation between the number of CLs and the number of viable and transferable embryos, but the number of CLs and the number of oocytes and/or degenerated embryos were not correlated. A total of 46 NsDU ETs were performed in nonhormonally treated recipient sows, with embryos (30 embryos per transfer) recovered from the 1000-IU eCG, 1500-IU eCG, and control groups. In total, pregnancy and farrowing rates were 75.1% and 73.2%, respectively, with a litter size of 9.4 ± 0.6 piglets born, of which 8.8 ± 0.5 were born alive. There were no differences for any of the reproductive parameters evaluated among groups. In conclusion, our results demonstrated the efficiency of eCG superovulation treatments in decreasing the donor-to-recipient ratio. Compared with nonsuperovulated sows, the number of transferable embryos was increased in superovulated sows without affecting their quality and in vivo capacity to develop to term after transfer. The results from this study also demonstrate the effectiveness of the NsDU ET procedure used, making possible the commercial use of ET technology by the pig industry.

Forskolin improves the cryosurvival of in vivo-derived porcine embryos at very early stages using two vitrification methods.

This study was aimed to determine the effect of forskolin on the viability of in vivo-derived porcine embryos vitrified by the superfine open pulled straw (SOPS) or solid surface vitrification (SSV) methods at the 2-cell, 4-cell, and blastocyst stages. Zygotes, 2- to 4-cell embryos, and morulae were obtained from superovulated sows. After collection, embryos were cultured for 24h with 0 or 10 µM forskolin and then vitrified using the SOPS and SSV method, or not vitrified (fresh controls). Fresh and vitrified-warmed 2-cells, 4-cells, and blastocysts were cultured for additional 96 h, 72 h and 24 h, respectively. At the end of the culture, embryos were evaluated for progression to the blastocyst stage and total cell number. The vitrification method did not affect any of the parameters evaluated for any embryo stage. Forskolin increased (P<0.01) the blastocyst formation and the final developmental stage of vitrified 2- and 4-cell embryos. However, these embryos exhibited lower (P<0.003) blastocyst formation rates than their fresh counterparts. The total cell number and hatching rate were similar in both groups (vitrified and fresh) of 2- and 4-cell embryos. Vitrified blastocysts exhibited viabilities, final developmental stages, hatching rates, and total cell numbers that were similar to those of their fresh counterparts, regardless of the addition of forskolin. In conclusion, the SOPS and SSV methods are suitable for the cryopreservation of in vivo-derived 2- to 4-cell porcine embryos. Pre-treatment with forskolin for 24h before vitrification improves the cryotolerance of 2- and 4-cell porcine embryos.

Effects of lipid polarisation on survival of in vivo-derived porcine zygotes vitrified by the superfine open pulled-straw method.

This study aimed to evaluate the post-warming in vitro viability of intact porcine zygotes vitrified using the superfine open pulled-straw (SOPS) method and to investigate whether cryotolerance is increased by lipid polarisation before vitrification. In vivo-derived zygotes (n=317) were either untreated before SOPS vitrification or subjected to one of the following pre-treatments: (1) centrifugation (20 min, 15000 g) or (2) equilibration in high-osmolality medium (6 min, 400 mOsm kg(-1)) followed by centrifugation. Vitrified-warmed and non-vitrified fresh zygotes were cultured in vitro for 120 h. There were no differences in the blastocyst formation rates between the vitrification groups (from 35.4±5.3% to 48.2±5.6%), but fresh zygotes exhibited higher (P<0.001) blastocyst formation rates (87.5±5.3%) than did vitrified-warmed zygotes. The total blastocyst cell number was similar among all groups (from 34.9±2.8 to 44.1±2.8). In conclusion, SOPS vitrification is a promising method for the cryopreservation of untreated in vivo-derived porcine zygotes. Neither lipid polarisation by centrifugation nor exposure to a high-osmolality medium followed by centrifugation affected the post-warming in vitro viability of zygotes. Our study also demonstrated that the donor is an important factor in determining the success of vitrification for in vivo-derived porcine zygotes.

The in vitro and in vivo developmental capacity of selected porcine monospermic zygotes.

In this study, we evaluated the in vitro and in vivo developmental capacity of selected monospermic zygotes produced in vitro. Cumulus-oocyte complexes were matured in vitro and inseminated with frozen-thawed spermatozoa. Thirteen hours after insemination, presumptive zygotes were centrifuged at 15,000 ×g for 20 minutes to polarize the lipids in the cytoplasm and permit the visualization of pronuclei. Then, the oocytes were individually classified as bipronuclear (2PN) or polypronuclear (three or more pronuclei, PPN). To examine embryo development, 102 selected zygotes were cultured for 7 days. There were no differences in cleavage rate (93.0% and 88.9% for 2PN and PPN zygotes, respectively). However, the blastocyst formation rate was higher (P < 0.003) in 2PN (80.7%) zygotes than in PPN (53.3%) zygotes. The control (noncentrifuged, nonselected zygotes) group showed lower (P < 0.003) cleavage rate and blastocyst formation than the 2PN and PPN zygotes. In a second experiment, 2PN zygotes and control zygotes were transferred (30 zygotes per transfer) by laparoscopy into the oviducts of recipient gilts (10 recipients per group) on the first day of standing estrus. The farrowing rates were 70% and 40% for transfers made with 2PN and control zygotes, respectively. The average number of piglets born per recipient farrowed did not differ between groups (4.9 ± 0.6 and 4.5 ± 1.2, respectively), but the efficiency (number of live piglets per total transferred embryos) was higher (P < 0.01) for 2PN zygotes than for the control group (9.3% and 4.0%, respectively). These results demonstrate the effectiveness of centrifugation for the selection of monospermic zygotes as a procedure to improve in vitro embryo production in pigs. In addition, the results indicate that the laparoscopic technique described here is a simple and effective procedure for transferring embryos into one oviduct.

Effect of MEM vitamins and forskolin on embryo development and vitrification tolerance of in vitro-produced pig embryos.

The aims of this study were (1) to determine the effect of in vitro maturation (IVM) medium supplementation with MEM vitamins on in vitro embryo development and sensitivity to vitrification of Day 6 blastocysts and (2) to evaluate whether the addition of forskolin to in vitro culture (IVC) medium enhances blastocyst survival following Super Open Pulled Straw (SOPS) vitrification. Cumulus-oocyte complexes (COCs; n=4000) were matured with 0.0% or 0.05% (v/v) MEM vitamins. After 44h of IVM, the oocytes were in vitro fertilized, and presumptive zygotes were cultured. At Day 5 of IVC, embryos from both experimental groups were cultured for 24h with 0 or 10µM forskolin, achieving a 2×2 factorial design. The blastocyst formation rate was assessed on Day 6, and subsets of samples from the four experimental groups were vitrified (n=469) or kept fresh (n=546). Fresh and vitrified-warmed blastocysts were cultured for 24h prior to embryo survival and total blastocyst cell number assessment. The MEM vitamins increased (P<0.001) the blastocyst formation rate at Day 6, but they did not affect embryo survival after vitrification. In contrast, the addition of forskolin to the culture medium enhanced (P<0.05) the blastocyst vitrification tolerance. The total blastocyst cell number was similar among the groups. In conclusion, supplementation with 0.05% MEM vitamins improved the blastocyst formation rate, and the addition of 10µM forskolin to the culture medium increased survival in Day 6 in vitro-produced blastocysts after SOPS vitrification.

Non-surgical deep intrauterine transfer of superfine open pulled straw (SOPS)-vitrified porcine embryos: evaluation of critical steps of the procedure.

Previous trials achieved extremely poor results when using the one-step warming method in a syringe in combination with non-surgical deep intrauterine transfer (NET) of superfine open pulled straw (SOPS)-vitrified embryos. This study aimed to assess the effect of the warming procedure on the in vitro and in vivo development of SOPS-vitrified embryos. The effect of the passage of the vitrified-warmed (VW) embryos through the NET catheter was also evaluated. Groups of 4 to 6 morulae and blastocysts, collected from weaned sows, were SOPS-vitrified in 1 µL of vitrification medium, warmed by the one-step warming method in a dish or in a 1-mL syringe and cultured in vitro for 48 h to evaluate the embryo survival (ES) and hatching rates (HR). Warming in syringe had a deleterious effect (P < 0.05) on the in vitro ES (60.5 ± 10.4%) and HR (39.6 ± 9.5%) of VW embryos in comparison with embryos warmed in a dish (85.4 ± 10.6% and 69.0 ± 8.4%, respectively). This decreased embryonic development was due to the increased time required between the removal of the straws from the liquid nitrogen and the contact of the embryos with the warming medium when the warming was performed in a syringe in comparison with that for the warming in a dish. After verifying that the passage of VW embryos through the NET catheter does not have a damaging effect on their further in vitro development, the negative effect of warming in a syringe was also confirmed after NET. Fifteen fresh and SOPS-vitrified embryos warmed in a syringe or in a dish were transferred to each recipient (n = 28) and recovered 24 h later to assess their developmental progression. All embryos from the syringe group were found to have degenerated at recovery. The in vivo ES and HR from the dish group (80.4 ± 3.4% and 14.2 ± 7.2%, respectively) were lower (P < 0.05) than those from the fresh group (94.0 ± 4.1% and 36.8 ± 7.8%, respectively). Combining the warming in a dish and the NET procedure, 35 VW embryos were transferred to each of 10 gilts. Five recipients farrowed an average of 10.4 ± 0.9 piglets. In conclusion, the method of one-step warming in a syringe has a negative effect on the in vitro and in vivo viability of SOPS-vitrified porcine embryos. In addition, NET of SOPS-vitrified embryos warmed by the one-step method in a dish showed promising reproductive performance of recipients. However, despite the great potential of this technology, further developments are required for large-scale commercial applications.

[Prevalence of Helicobacter pylori detected by real-time PCR in the subgingival plaque of patients with chronic periodontitis]. / Prévalence d'helicobacter pylori détecté par PCR en temps réel dans la plaque sous-gingivale des patients atteints de parodontite chronique.

BACKGROUND: Helicobacter pylori plays a significant role in gastric disease. The aim of the present pilot study was to evaluate the prevalence of H. pylori detected by real-time polymerase chain reaction (real-time PCR) in the sub-gingival plaque of chronic periodontitis patients and to determine periodontopathogens profile of positive sites at H. pylori. METHODOLOGY: 109 subgingival samples collected from 17 subjects with chronic periodontitis were studied. The DNA was extracted from the oral samples and analyzed for the presence of H. pylori by real-time PCR (LightCycler) using 16S rRNA#120 primers which targeted the 16S rRNA gene. DNA from H. pylori DSM 4867 was used as a positive control. Seven bacteria implicated in chronic periodontitis were selected to explore the presence of these periodontopathic species in the oral positive sites for H. pylori. RESULTS: 16 of 109 samples (14.7%) were positives of H. pylori. All the positives sites were also positives to Fusobacterium nucleatum and Eikenella corrodens, 62.5% to Porphyromonas gingivalis, 31.25% to Treponema denticola, 25% to Prevotella intermedia, 12.5% to Aggregatibacter actinomycetemcomitans and 6.25% to Tannerella forsythia. CONCLUSIONS: H. pylori may be present in the subgingival plaque samples of patients with chronic periodontitis who are resident in a developing country. F. nucleatum and E. corrodens could coaggregate with H. pylori in the subgingival dental plaque.

Use of polarized light microscopy in porcine reproductive technologies.

The meiotic spindle in the oocyte is composed of microtubules and plays an important role during chromosome alignment and separation at meiosis. Polarized light microscopy (PLM) could be useful for a non-invasive evaluation of the meiotic spindle and may allow removal of nuclear structures without fluorochrome staining and ultraviolet exposure. In this study, PLM was used to assess its potential application in porcine reproductive technologies. The objectives of the present study were to assess the efficiency of PLM to detect microtubule-polymerized protein in in vitro-matured porcine oocytes; to examine its effects on the oocyte developmental competence; to select oocytes based on the presence of the meiotic spindle detected by PLM; and to assess the efficiency oocyte enucleation assisted with PLM. In the first experiment, the presence of microtubule-polymerized protein was assessed and confirmed in oocytes (n = 117) by immunostaining and chromatin detection. In the second experiment, oocytes (n = 160) were exposed or not (controls) to PLM for 10 minutes, and then parthenogenetically activated and cultured in vitro. In the third experiment, development competence of oocytes with a positive or negative signal to PLM was analyzed after in vitro fertilization. Finally, oocytes (n = 54) were enucleated using PLM as a tool to remove the meiotic spindle. A positive PLM signal was detected in 98.2 % of the oocytes, which strongly correlated (r = 1; p < 0.0001) with the presence of microtubule-polymerized protein as confirmed by immunostaining. Oocytes exposed to PLM did not differ significantly from controls on cleavage, total blastocyst, expanded blastocyst rates and total cell numbers. The percentage of oocytes at the MII stage and blastocyst formation rate in the negative PLM group significantly differed from control and PLM positive groups. Overall efficiency of spindle removal using the PLM-Oosight system was 92.6%. These results suggest that polarized light microscopy is an efficient system to detect microtubule-polymerized protein in in vitro-matured porcine oocytes and does not exert detrimental effects on porcine oocyte developmental competence. Selecting oocytes by the presence of a PLM signal provides limited improvement on IVF results. Finally, PLM appears as an efficient method to enucleate porcine oocytes.

[Comparison of four different primer sets for detection of Helicobacter pylori in gastric biopsies and oral samples by using real-time PCR]. / Comparaison de quatre paires d'amorces différentes dans la détection d'Helicobacter pylori dans les biopsies gastriques et les prélèvements oraux.

AIMS: The presence of Helicobacter pylori. in the oral cavity remains controversial and the most appropriate method for detection of oral H. pylori has yet to be established. The aim of the present study was to compare four different primer sets on the detection of H. pylori in gastric biopsies and oral samples using real-time PCR. MATERIAL AND METHODS: Gastric biopsy and oral samples were collected from eight patients with gastric symptoms. DNA from clinical samples was extracted and analyzed for the presence of H. pylori by real-time PCR (LightCycler 2.0) using four pairs of primers which targeted 16S rRNA (16S rRNA#295; 16S rRNA#120) or glmM (glmM#294; glmM#722) DNA genes. Three H. pylori strains and three clinical isolates served as reference. The specificities of the four primer pairs were examined for seven oral microorganisms and two Helicobacter non-pylori species. RESULTS: Primer pair 16S rRNA#120 showed an acceptable specificity and a high sensitivity. Primer pairs glmM#294 and glmM#722 demonstrated a high specificity but a low sensitivity and primer pair 16S rRNA#295 demonstrated a poor specificity but acceptable sensitivity. Four H. pylori positive gastric biopsies were demonstrated by culture, histology and real-time PCR with primer pairs 16S rRNA#295 or 16S rRNA#120. No H. pylori was detected in oral samples, either by culture or by real-time PCR. CONCLUSION: Of the four different primer pairs examined, 16S rRNA#120 was the most appropriate to detect H. pylori in clinical samples using real-time PCR.