How to identify patients who would benefit from delayed-matured oocytes insemination: a sibling oocyte and ploidy outcome study.

STUDY QUESTION: Which patients might benefit from insemination of delayed-matured oocytes? SUMMARY ANSWER: Delayed-matured oocytes had a ≥50% contribution to the available cohort of biopsied blastocysts in patients with advanced maternal age, low maturation, and/or low fertilization rates. WHAT IS KNOWN ALREADY: Retrieved immature oocytes that progress to the MII stage in vitro could increase the number of embryos available during ICSI cycles. However, these delayed-matured oocytes are associated with lower fertilization rates and compromised embryo quality. Data on the ploidy of these embryos are controversial, but studies failed to compare euploidy rates of embryos derived from delayed-matured oocytes to patients' own immediate mature sibling oocytes. This strategy efficiently allows to identify the patient population that would benefit from this approach. STUDY DESIGN, SIZE, DURATION: This observational study was performed between January 2019 and June 2021 including a total of 5449 cumulus oocytes complexes from 469 ovarian stimulation cycles, from which 3455 inseminated matured oocytes from ICSI (n = 2911) and IVF (n = 544) were considered as the sibling controls (MII-D0) to the delayed-matured oocytes (MII-D1) (n = 910). Euploidy rates were assessed between delayed-matured (MII-D1) and mature sibling oocytes (MII-D0) in relation to patients' clinical characteristics such as BMI, AMH, age, sperm origin, and the laboratory outcomes, maturation, fertilization, and blastocyst utilization rates. PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 390 patients undergoing IVF/ICSI, who had at least one metaphase I (MI) or germinal-vesicle (GV) oocyte on the day of oocyte collection (Day 0), which matured in 20-28 h after denudation were included. MI and GV oocytes that matured overnight were inseminated on the following day (Day 1, MII-D1) by ICSI. Only cycles planned for preimplantation genetic testing for aneuploidy using fresh own oocytes were included. MAIN RESULTS AND THE ROLE OF CHANCE: Fertilization (FR) and blastocyst utilization rates were significantly higher for MII-D0 compared to delayed-matured oocytes (MII-D1) (69.5% versus 55.9%, P < 0.001; and 59.5% versus 18.5%, P < 0.001, respectively). However, no significant difference was observed in the rate of euploid embryos between MII-D0 and MII-D1 (46.3% versus 39.0%, P = 0.163). For evaluation of the benefit of inseminating MI/GV oocytes on D1 per cycle in relation to the total number of biopsied embryos, cycles were split into three groups based on the proportion of MII-D1 embryos that were biopsied in that cycle (0%, 1-50%, and ≥50%). The results demonstrate that patients who had ≥50% contribution of delayed-matured oocytes to the available cohort of biopsied embryos were those of advanced maternal age (mean age 37.7 years), <10 oocytes retrieved presenting <34% maturation rate, and <60% fertilization rate. Every MII oocyte injected next day significantly increased the chances of obtaining a euploid embryo [odds ratio (OR) = 1.83, CI: 1.50-2.24, P < 0.001] among MII-D1. The odds of enhanced euploidy were slightly higher among the MII-D1-GV matured group (OR = 1.78, CI: 1.42-2.22, P < 0.001) than the MII-D1-MI matured group (OR = 1.54, CI: 1.25-1.89, P < 0.001). Inseminating at least eight MII-D1 would have >50% probability of getting a euploid embryo among the MII-D1 group. LIMITATIONS, REASONS FOR CAUTION: ICSI of MII-D1 was performed with the fresh or frozen ejaculates or testicular samples from the previous day. The exact timing of polar body extrusion of delayed-matured MI/GV was not identified. Furthermore, the time point of the final oocyte maturation to MII for the immature oocytes and for the oocytes inseminated by IVF could not be identified. WIDER IMPLICATIONS OF THE FINDINGS: The results of this study might provide guidance to the IVF laboratories for targeting the patient's population who would benefit from MII-D1 ICSI without adhering to unnecessary costs and workload. STUDY FUNDING/COMPETING INTEREST(S): No external funding was received for this study. There are no conflicts of interest to be declared for any of the authors. There are no patents, products in development, or marketed products to declare. TRIAL REGISTRATION NUMBER: N/A.

A hybrid artificial intelligence model leverages multi-centric clinical data to improve fetal heart rate pregnancy prediction across time-lapse systems.

STUDY QUESTION: Can artificial intelligence (AI) algorithms developed to assist embryologists in evaluating embryo morphokinetics be enriched with multi-centric clinical data to better predict clinical pregnancy outcome? SUMMARY ANSWER: Training algorithms on multi-centric clinical data significantly increased AUC compared to algorithms that only analyzed the time-lapse system (TLS) videos. WHAT IS KNOWN ALREADY: Several AI-based algorithms have been developed to predict pregnancy, most of them based only on analysis of the time-lapse recording of embryo development. It remains unclear, however, whether considering numerous clinical features can improve the predictive performances of time-lapse based embryo evaluation. STUDY DESIGN, SIZE, DURATION: A dataset of 9986 embryos (95.60% known clinical pregnancy outcome, 32.47% frozen transfers) from 5226 patients from 14 European fertility centers (in two countries) recorded with three different TLS was used to train and validate the algorithms. A total of 31 clinical factors were collected. A separate test set (447 videos) was used to compare performances between embryologists and the algorithm. PARTICIPANTS/MATERIALS, SETTING, METHODS: Clinical pregnancy (defined as a pregnancy leading to a fetal heartbeat) outcome was first predicted using a 3D convolutional neural network that analyzed videos of the embryonic development up to 2 or 3 days of development (33% of the database) or up to 5 or 6 days of development (67% of the database). The output video score was then fed as input alongside clinical features to a gradient boosting algorithm that generated a second score corresponding to the hybrid model. AUC was computed across 7-fold of the validation dataset for both models. These predictions were compared to those of 13 senior embryologists made on the test dataset. MAIN RESULTS AND THE ROLE OF CHANCE: The average AUC of the hybrid model across all 7-fold was significantly higher than that of the video model (0.727 versus 0.684, respectively, P = 0.015; Wilcoxon test). A SHapley Additive exPlanations (SHAP) analysis of the hybrid model showed that the six first most important features to predict pregnancy were morphokinetics of the embryo (video score), oocyte age, total gonadotrophin dose intake, number of embryos generated, number of oocytes retrieved, and endometrium thickness. The hybrid model was shown to be superior to embryologists with respect to different metrics, including the balanced accuracy (P ≤ 0.003; Wilcoxon test). The likelihood of pregnancy was linearly linked to the hybrid score, with increasing odds ratio (maximum P-value = 0.001), demonstrating the ranking capacity of the model. Training individual hybrid models did not improve predictive performance. A clinic hold-out experiment was conducted and resulted in AUCs ranging between 0.63 and 0.73. Performance of the hybrid model did not vary between TLS or between subgroups of embryos transferred at different days of embryonic development. The hybrid model did fare better for patients older than 35 years (P < 0.001; Mann-Whitney test), and for fresh transfers (P < 0.001; Mann-Whitney test). LIMITATIONS, REASONS FOR CAUTION: Participant centers were located in two countries, thus limiting the generalization of our conclusion to wider subpopulations of patients. Not all clinical features were available for all embryos, thus limiting the performances of the hybrid model in some instances. WIDER IMPLICATIONS OF THE FINDINGS: Our study suggests that considering clinical data improves pregnancy predictive performances and that there is no need to retrain algorithms at the clinic level unless they follow strikingly different practices. This study characterizes a versatile AI algorithm with similar performance on different time-lapse microscopes and on embryos transferred at different development stages. It can also help with patients of different ages and protocols used but with varying performances, presumably because the task of predicting fetal heartbeat becomes more or less hard depending on the clinical context. This AI model can be made widely available and can help embryologists in a wide range of clinical scenarios to standardize their practices. STUDY FUNDING/COMPETING INTEREST(S): Funding for the study was provided by ImVitro with grant funding received in part from BPIFrance (Bourse French Tech Emergence (DOS0106572/00), Paris Innovation Amorçage (DOS0132841/00), and Aide au Développement DeepTech (DOS0152872/00)). A.B.-C. is a co-owner of, and holds stocks in, ImVitro SAS. A.B.-C. and F.D.M. hold a patent for 'Devices and processes for machine learning prediction of in vitro fertilization' (EP20305914.2). A.D., N.D., M.M.F., and F.D.M. are or have been employees of ImVitro and have been granted stock options. X.P.-V. has been paid as a consultant to ImVitro and has been granted stocks options of ImVitro. L.C.-D. and C.G.-S. have undertaken paid consultancy for ImVitro SAS. The remaining authors have no conflicts to declare. TRIAL REGISTRATION NUMBER: N/A.

Bone volume-to-total volume ratio measured in trabecular bone by single-sided NMR devices.

PURPOSE: Reduced bone strength is associated with a loss of bone mass, usually evaluated by dual-energy X-ray absorptiometry, although it is known that the bone microstructure also affects the bone strength. Here, a method is proposed to measure (in laboratory) the bone volume-to-total volume ratio by single-sided NMR scanners, which is related to the microstructure of the trabecular bone. METHODS: Three single-sided scanners were used on animal bone samples. These low-field, mobile, low-cost devices are able to detect the NMR signal, regardless of the sample sizes, without the use of ionizing radiations, with the further advantage of signal localization offered by their intrinsic magnetic field gradients. RESULTS: The performance of the different single-sided scanners have been discussed. The results have been compared with bone volume-to-total volume ratio by micro CT and MRI, obtaining consistent values. CONCLUSIONS: Our results demonstrate the feasibility of the method for laboratory analyses, which are useful for measurements like porosity on bone specimens. This can be considered as the first step to develop an NMR method based on the use of a mobile single-sided device, for the diagnosis of osteoporosis, through the acquisition of the signal from the appendicular skeleton, allowing for low-cost, wide screening campaigns. Magn Reson Med 79:501-510, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

M2 macrophages coexist with a Th1-driven profile in periapical cysts.

AIM: To evaluate the participation of both Th1 and Th2 responses in periapical cysts by assessing the presence of M2 macrophages, as well as acute IL-1 ß, TNF-α and IL-6 cytokines. METHODOLOGY: Twenty-four cases of periapical cysts were selected. Immuno-expressions of IL-1 ß, IL-6, TNF-α and CD163 were analysed in the cystic capsules in both superficial and deeper regions. Data were analysed with paired Wilcoxon test and Spearman correlation coefficient (P ≤ 0.05). RESULTS: There was a higher expression of IL-1ß, IL-6, TNF-α and M2 macrophages in the superficial region (P < 0.001) of cystic capsules. All acute cytokines had significant positive correlations amongst them regardless of the cystic capsule region. Regarding CD163, positive correlations occurred only with TNF-α (P = 0.007; r = 0.537) and IL-6 (P = 0.018; r = 0.478) in the superficial regions of the cystic capsule. CONCLUSIONS: M2 macrophages participated actively in the inflammatory response of periapical cysts and correlated with the expression of certain acute Th1-related cytokines. This illustrates the coexistence of an acute and chronic Th2-driven immune response in these lesions. Although M2 macrophages favour the healing process, their presence is not sufficient for periapical cyst regression, once an acute active response has occurred due to an infectious stimuli.

Quercetin influences in vitro maturation, apoptosis and metabolically active mitochondria of goat oocytes.

SummaryThe present study aimed to investigate the effect of quercetin as an alternative antioxidant to cysteamine on in vitro maturation. Oocytes were collected from goat ovaries, destined for in vitro maturation and distributed into three groups: CIS group, oocytes were immersed in MIV base medium; in Groups Q4 and Q8, oocytes were immersed in the medium of the CIS group, adding 4 µM or 8 µM of quercetin, respectively, and cultured for 24 h at 38.5°C with 5% CO2. The CIS and Q4 groups presented the same percentage of expanded cumulus cells, but the per cent in the Q8 group was significantly lower than that of the other groups (P<0.05). The oocyte retraction rate in the Q8 group was higher (P<0.05) than in the CIS and Q4 groups. Treatment with 8 µM of quercetin presented a lower proportion of expanded oocytes than the CIS group and 4 µM of quercetin (P<0.05). The percentage of MII oocytes was higher in the Q4 group than in the CIS group (P<0.05), but the percentages in the CIS and Q8 groups were similar. The rate of apoptosis was higher in the CIS group than in the other groups (P<0.05). In addition, oocytes matured with 4 µM quercetin showed higher mitochondrial activity than matured oocytes in the CIS and Q8 groups (P<0.05). In conclusion, 4 µM of quercetin can be used as an alternative to cysteamine in the in vitro maturation of goat oocytes.

Natural products are the future of anticancer therapy: Preclinical and clinical advancements of Viscum album phytometabolites.

Cancer is a multifaceted and genomically complex disease. Research over the years has gradually provided a near complete resolution of cancer landscape and it is now known that genetic/epigenetic mutations, inactivation of tumor suppressors, Overexpression of oncogenes, spatio-temporally dysregulated intracellular signaling cascades, epithelial to mesenchymal transition (EMT), metastasis and loss of apoptosis are some of the most extensively studied biological mechanisms that underpin cancer development and progression. Increasingly it is being realized that current therapeutic interventions are becoming ineffective because of tumor heterogeneity and rapidly developing resistance against drugs. Considerable biological activities exerted by bioactive ingredients isolated from natural sources have revolutionized the field of natural product chemistry and rapid developments in preclinical studies are encouraging. Viscum album has emerged as a deeply studied natural source with substantial and multifaceted biological activities. In this review we have attempted to provide recent breakthroughs in existing scientific literature with emphasis on targeting of protein network in cancer cells. We partition this review into different sections, highlighting latest information from cell culture studies, preclinical and clinically oriented studies. We summarized how bioactive ingredients of Viscum album modulated extrinsic and intrinsic pathways in cancer cells. However, surprisingly, none of the study reported stimulatory effects on TRAIL receptors. The review provided in-depth analysis of how Viscum album modulated Endoplasmic Reticulum Stress in cancer cells and how bioactive chemicals tactfully targeted cytoskeletal machinery in cancer cells as evidenced by cell culture studies. It is noteworthy that Viscum album has entered into various phases of clinical trials, however, there are still knowledge gaps in our understanding regarding how various bioactive constituents of Viscum album modulate intracellular signaling cascades in cancer. Better and deeper comprehension oncogenic signaling cascades will prove to be helful in getting a step closer to individualized medicine.

Selection indices to identify drought-tolerant grain sorghum cultivars.

Twenty-five cultivars of grain sorghum [Sorghum bicolor (L.) Moench] were examined under both drought stress and normal conditions in 4 experiments. In each condition, genotypes were evaluated in a factorial experiment using a randomized complete block design with 3 replications. Eight drought tolerance indices including stability tolerance index, mean productivity (MP), geometric MP, harmonic mean, stress susceptibility index, tolerance index, yield index, and yield stability index were estimated for each genotype based on grain yield under drought (Ys) and irrigated conditions (Yp). The results indicated that there were positive and significant correlations among Yp and Ys with geometric MP, MP, harmonic mean, and stability tolerance index, indicating that these factors are better predictors of Yp and Ys than tolerance index, stress susceptibility index, yield stability index, and yield index. Based on adjusted means at Yp and Ys, indices geometric MP, MP, harmonic mean, and stability tolerance index, unweighted pair group method with arithmetic mean cluster and biplot analysis, the most tolerant cultivars were '9929020', '9929034', and 'N 95B'.

Anthelmintic intoxication in goats and sheep: A systematic review.

The objective of this study was to conduct research of the literature available in electronic media on anthelmintic intoxication in sheep and goats. The search for primary studies was carried out in five electronic databases: ScienceDirect, PubMed, Scopus, Web of Science, and SciELO. The search terms used were (antihelmintic OR antiparasitic OR vermifuge) AND (poisoning OR toxicity OR overdose OR intoxication) AND (goat OR sheep). A total of 2361 articles were identified from the five databases: Science Direct (n = 1869), PubMed (n = 434), Scopus (n = 37), Web of Science (n = 16), and SciELO (n = 5). As 111 articles were found in duplicates, 2250 were left for review of the title and abstracts, of which 115 were read in full, and 28 were included in the systematic review. Of the 28 articles, 16 involved sheep, 9 involved goats, and 3 involved both species. Twelve drugs were identified in intoxication reports: albendazole (2), closantel (14), disophenol (1), ivermectin (1), levamisole (2), moxidectin (1), netobimin (1), nitroxinil (1), oxfendazole (2), parbendazole (2), tetramizole (1), and thiabendazole (1). The most prevalent symptoms of anthelmintic intoxication reported were showed involvement of the nervous, locomotor, and renal systems, as well as teratogenic influences. Data from this review underscore the need of the care required in the control of parasitic infections through the safe use of antiparasitic drugs to avoid cases of intoxication.

A New Species of Tetragona Lepeletier & Serville, 1828 from the "truncata group" and New Distribution Records of T. truncata Moure, 1971 (Hymenoptera: Apidae).

Tetragona Lepeletier & Serville is a genus of stingless bees with 14 recognized species occurring from Mexico to Argentina. The genus is characterized by velvety genal area, mesotibial spur present, and propodeal triangle glabrous. Within the genus, the truncata species group (T. truncata Moure and T. atahualpa sp. nov.) is characterized by worker metabasitarsus with posterior angle rounded and the mandible with two short teeth of similar length. Tetragona truncata is reported with new records for Ecuador (Napo and Orellana), Peru (Huánuco, Loreto, and San Martín), and Brazil (Acre [Rio Branco] and Tocantins [Itacá, Lizarda and Palmas]). In addition, T. atahualpa sp. nov. is described as a new species from regions of altitudes above 1,800 m in Colombia (Boyacá), Ecuador (Napo, Zamora-Chinchipe), and Peru (Pasco). We illustrate and discuss the identification of these two species.

Anabolic steroids and their effects of on neuronal density in cortical areas and hippocampus of mice.

Anabolic substances have been increasingly used by bodybuilders and athletes with the goal of improving performance and aesthetics. However, this practice has caused some concern to physicians and researchers because of unknowledge of consequences that the indiscriminate and illicit use of these substances can cause. Thus, this study analyzed the effects of two commercially available anabolic steroids (AS), Winstrol Depot® (Stanozolol) and Deposteron® (Testosterone Cypionate), in the neuronal density of limbic, motor and sensory regions on the cerebral cortex and in CA1, CA2, CA3 regions of the hippocampus. A total of 60 Swiss mice were used (30 males and 30 females), separated into three groups: control and two experimental groups, which received the AAS. From each brain, homotypic and semi-serial samples were taken in frontal sections from areas established for the study. The results showed that females treated with testosterone cypionate presented a reduction in all regions tested and the ones treated with Stanozolol showed a decrease in some hippocampal areas. Regarding male animals, stanozolol led to a decrease in neuron number in one hippocampal region. These data allow us to conclude that supra-physiological doses of steroids used in this study, can cause considerable damage to nervous tissue with ultrastructural and consequently behavioral impairment. These changes could interfere with the loss of physical yield and performance of athletes and non-athletes and may cause irreparable damage to individuals making irresponsible use of anabolic steroids.

First Report of Monosporascus cannonballus on Watermelon in Brazil.

In 2008 and 2009, vine decline symptoms were observed in three watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) fields located in the municipalities of Mossoró (Rio Grande do Norte State) and Quixeré (Ceará State) in northeastern Brazil. Symptoms included yellowing of crown leaves just prior to harvest and collapse of many of the vines. Mean maximum daily temperatures for the first and second half of the season were 28.6 and 25.1°C, respectively. Affected plants exhibited necrotic root systems and lacked most of the secondary and tertiary feeder roots. Numerous perithecia on the roots contained asci and ascospores characteristics of Monosporascus cannonballus Pollack & Uecker (1,2). Small pieces of primary and secondary roots were surface disinfected and plated onto potato dextrose agar (PDA) medium with 0.5 g liter-1 of streptomycin sulfate and incubated for 7 days at 25°C in the dark. Hyphal tips from all colonies were transferred to PDA and further incubated for 30 to 40 days at 25°C in the dark for subsequent growth and sporulation. Isolations consistently yielded colonies of white mycelium, which became dark grayish after 10 to 15 days, and perithecia with one-spored asci. The internal transcribed spacer regions of ribosomal DNA of isolates 18-5 and 19-1 were sequenced (GenBank Accession Nos. GQ891544 and GQ891545). These sequences were identical to sequences of M. cannonballus (GenBank Accession Nos. AM167936 and AM167937). Pathogenicity of these two isolates was confirmed on watermelon cv. Crimson Sweet in a greenhouse maintained at 25 to 30°C. Inoculum was produced in a sand-oat hulls (Avena sativa) medium (0.5 liter of sand, 46 g of ground oat hulls, and 37.5 ml of distilled water) and incubated at 25°C for 1 month. CFU were quantified by serial dilution using 1% hydroxyethyl cellulose. A sterilized mixture of equal portions (vol/vol) of sand and peat moss was used to fill 17-cm-diameter plastic pots and inoculum was added to produce an inoculum concentration of 20 CFU g-1. Five watermelon seeds planted in each pot were later thinned to one seedling per pot. There were five replicated pots for each treatment with an equal number of noninfested pots. Plants were evaluated for disease 45 days after sowing. All isolates of M. cannonballus were highly aggressive and caused severe root necrosis compared with the noninoculated controls. M. cannonballus was reisolated from symptomatic plants, confirming Koch's postulates. In 2004, M. cannonballus was reported in the same Brazilian cucurbit-growing areas causing root rot and vine decline of muskmelon (Cucumis melo L.) (3), but to our knowledge, this is the first report of M. cannonballus on watermelon in Brazil. References: (1) R. D. Martyn and M. E. Miller. Plant Dis. 80:716, 1996. (2) F. G. Pollack and F. A. Uecker. Mycologia 66:346, 1974. (3) R. Sales Jr. et al. Plant Dis. 88:84, 2004.

Plant extracts as natural modulators of gut microbiota community structure and functionality.

The main objective of this work was to evaluate the effect that several plant extracts (currently sold as functional ingredients) have on gut microbiota community structure and functionality. Plant extracts were submitted to an in vitro digestion and fecal fermentation. Overall, plant extracts showed a marked inhibitory activity when compared to basal conditions. However, they also favored the growth of some bacteria such as Coprococcus and Butyricimonas, two butyrate producers. Especially interesting was tea extract which inhibited the growth of the genus Escherichia/Shigella, known to involve species related with gastrointestinal disorders. Additionally, tea extract increased the growth of Faecalibacterium, a known butyrate producer. Regarding short chain fatty acids production, while plant extracts reduced acetate production, butyrate was increased for most samples, especially tea extract. Propionate production was less affected in comparison with basal conditions. Fermentation by gut microbiota also modified the antioxidant capacity (assessed via DPPH, FRAP and Folin-Ciocalteu methods).

Immunotherapy against Systemic Fungal Infections Based on Monoclonal Antibodies.

: The increasing incidence in systemic fungal infections in humans has increased focus for the development of fungal vaccines and use of monoclonal antibodies. Invasive mycoses are generally difficult to treat, as most occur in vulnerable individuals, with compromised innate and adaptive immune responses. Mortality rates in the setting of our current antifungal drugs remain excessively high. Moreover, systemic mycoses require prolonged durations of antifungal treatment and side effects frequently occur, particularly drug-induced liver and/or kidney injury. The use of monoclonal antibodies with or without concomitant administration of antifungal drugs emerges as a potentially efficient treatment modality to improve outcomes and reduce chemotherapy toxicities. In this review, we focus on the use of monoclonal antibodies with experimental evidence on the reduction of fungal burden and prolongation of survival in in vivo disease models. Presently, there are no licensed monoclonal antibodies for use in the treatment of systemic mycoses, although the potential of such a vaccine is very high as indicated by the substantial promising results from several experimental models.

Paracoccidioides HSP90 Can Be Found in the Cell Surface and Is a Target for Antibodies with Therapeutic Potential.

Paracoccidioidomycosis (PCM) is one of the most frequent systemic mycoses in Latin America. It affects mainly male rural workers in impoverished regions, and the therapy can last up to two years or use drugs that are very toxic. Given the need for novel safe and effective approaches to treat PCM, we have been developing monoclonal antibodies (mAbs) that could be used not only to block specific fungal targets, but also modulate the host's antifungal immunity. In this work we show the generation of and promising results with an mAb against Heat Shock Protein (HSP)90, a molecular chaperone that is an important virulence factor in fungi. Using recombinant Paracoccidioides lutzii (Pb01) and P. brasiliensis (Pb18) HSP90 proteins produced in E. coli, we immunized mice and generated polyclonal antibodies and an IgG1 hybridoma mAb. The proteins were very immunogenic and both the polyclonal serum and mAb were used in immunofluorescence experiments, which showed binding of antibodies to the yeast cell surface. The mAb successfully opsonized P. lutzii and P. brasiliensis cells in co-incubations with J774.16 macrophage-like cells. Our results suggest that this mAb could serve as the basis for new immunotherapy regimens for PCM.

Prematuration of human denuded oocytes in a three-dimensional co-culture system: effects on meiosis progression and developmental competence.

BACKGROUND: During conventional in vitro maturation (IVM), oocytes undergo nuclear maturation but do not attain full cytoplasmic maturity. The present study aimed to evaluate the quality of denuded in vitro matured human oocytes (= 'rescue IVM') following a prematuration culture (PMC) in a three-dimensional (3D) co-culture system with cumulus cells. METHODS: Denuded, germinal vesicle (GV) oocytes retrieved from controlled ovarian hyperstimulation (COH) cycles were embedded in collagen (Type I) gel containing dissociated cumulus cells and a specific phosphodiesterase 3-inhibitor (PDE3-I) for 24 h. Afterwards, oocytes were removed from the gel and transferred to IVM inhibitor-free medium. The reversibility of PDE3 inhibition was assessed following IVM culture for a maximum of 48 h. Cytoplasmic maturation was assessed by the oocytes' capability to fertilize after ICSI and sustain further embryonic development. Conventional microdroplet cultures of denuded oocytes, with and without PMC, served as controls. RESULTS: Oocytes embedded in collagen gel and exposed to PDE3-I were efficiently arrested at the GV stage. After removal from collagen and inhibitor, oocytes were capable of resuming meiosis: maturation rates (74.5%) were similar compared with non-prematured (68.5%) and prematured oocytes (73.4%) in microdroplets. Fertilization rates after PMC in 3D co-culture were significantly higher compared with non-prematured oocytes (72.2% versus 54.0%; P < 0.05), but similar to oocytes prematured in microdroplets (63.8%). Compared with both controls, the PMC in 3D co-culture improved the morphological quality and nuclear constitution of embryos on Day 3 post-ICSI. CONCLUSIONS: Three-dimensional PMC co-culture using a PDE3-I may enhance cytoplasmic maturation of denuded human GV oocytes retrieved from COH cycles.

Assessment of a new in vitro maturation system for mouse and human cumulus-enclosed oocytes: three-dimensional prematuration culture in the presence of a phosphodiesterase 3-inhibitor.

BACKGROUND: Controlling nuclear maturation during oocyte culture might improve nuclear-cytoplasmic maturation synchrony. In the present study, the quality of mouse and human cumulus-enclosed oocytes (CEOs) was examined after a two-step culture consisting of a three-dimensional prematuration culture (3D-PMC), followed by in vitro maturation (IVM). METHODS: Mouse and human CEOs were embedded in an extracellular matrix (collagen-gel Type I). The gels containing the CEOs were cultured in medium with a phosphodiesterase 3-inhibitor (PDE3-I; cilostamide 1 microM) for 24 h. Afterwards, CEOs were removed from the gel and washed away from inhibitor then underwent IVM. The optimal concentration of collagen (diluted 1:2 versus not-diluted) was first determined in the mouse model. Cytoplasmic maturation after IVM of human and mouse oocytes was assessed in relation to fertilization and embryonic developmental capacity. RESULTS: The diluted form of collagen was better for supporting the structure of the expanding CEOs and meiotic competence of the oocytes. Electron microscopy in combination with Lucifer Yellow dye coupling assay revealed that oocyte-cumulus cell connections could be preserved during 3D-PMC. Percentages of mouse 2-cell embryos after IVF were higher in the 3D-PMC group compared with in vitro controls and 2D-PMC oocytes, but lower compared with in vivo controls. In the human model, percentages of polar body-extruded oocytes were significantly higher in the 3D-PMC group compared with conventionally matured oocytes. The 3D-PMC also had a beneficial effect on embryonic development on Day 3 post-ICSI. CONCLUSIONS: Applying a 3D-PMC in the presence of a PDE3-I preserves oocyte-cumulus cell connections and influences oocyte developmental capacity.

Qafzeh 9 mandible (ca 90-100 kyrs BP, Israel) revisited: µ-CT and 3D reveal new pathological conditions.

OBJECTIVE: The aim of this paper is to provide new insights into growth patterns and health of Mousterian hunter-gatherers dated to ca. 90-100 kyrs B.P. from the Qafzeh site. MATERIALS: An almost complete skeleton, including the mandible from the Qafzeh site (Qafzeh 9). METHODS: Micro-CT and medical imaging techniques are used to explore inaccessible inner structures and to assess the etiology of identified lesion. RESULTS: Mandibular and dental conditions appear to be growth-related skeletal disorders. CONCLUSION: To our knowledge, Qafzeh 9 offers the earliest evidence of associated mandibular and dental pathological conditions (i.e. non-ossifying fibroma of the mandible, pre-eruptive intracoronal resorption and osteochondritis dissecans of the temporomandibular joint) among early anatomically modern humans, and more generally among Middle Palaeolithic hominins in Southwestern Asia. The diagnoses can be added to other growth-related disorders of skulls previously documented from the Qafzeh site (Tillier, 1999; Tillier et al., 2001), suggesting a quite high and exceptional incidence of these conditions compared to those of Palaeolithic populations. SIGNIFICANCE: Identification of pathological conditions in subadults from Eurasia dated to the Middle Palaeolithic is documented less often than in adult skeletons. Hence, new results from immature fossil remains are particularly important to our understanding of the past. SUGGESTIONS FOR FURTHER RESEARCH: This research will be extended to the rest of the skeleton, then to all Qafzeh specimens in order to broaden our understanding of this anatomically modern group.

Environmental Thresholds of Nepomorpha in Cerrado Streams, Brazilian Savannah.

Determining thresholds of species loss in response to gradients of human impact is becoming an increasingly important component of stream conservation and management. We analyzed the thresholds of change in populations and communities of the infraorder Nepomorpha along a gradient of environmental integrity in streams of the Brazilian savannah, based on samples from 20 sites. We used the Threshold Indicator Taxa Analysis (TITAN) to identify the location and magnitude of changes associated with the environmental quality and the frequency and abundance of the different taxa. We also used TITAN to calculate the positive and negative thresholds of the community and species along the habitat integrity gradient. Nepomorphan species were generally associated with sites of higher environmental quality, which presented larger populations and greater species richness. Only a few nepomorphan species groups were associated with impacted streams. Thresholds were evident in some aquatic nepomorphan populations, but not for the community as a whole. This indicates that species have idiosyncratic responses to the environmental gradient, but in general nepomorphans appear to be a good indicator of healthy environments or less impacted streams in the Brazilian savannah.

[Blastocyst quality criteria]. / Critères de qualité des blastocystes.

Embryo culture to the blastocyst stage has progressed enormously with the new generation of acellular culture media, or sequential media. The main advantages of embryo transfer at the blastocyst stage are that it provides a natural embryo selection during culture, or a selection using preimplantation diagnosis, it provides better conditions for SET, avoiding multiple pregnancies and overcoming repeated IVF failures by improving embryo selection. The risks are failing to select a single blastocyst or obtaining a limited number of frozen blastocysts, with reduced survival after thawing. However, prolonged embryo culture seems to be good practice for day 3 embryos with delayed development, and blastocyst transfer. More effective at day 5 than at day 6.