Effect of sperm treatment with lysolecithin on in vitro outcomes of equine intracytoplasmic sperm injection.

Intracytoplasmic sperm injection (ICSI) in horses is currently employed for clinical and commercial uses, but the protocol could be optimized to improve its efficiency. We have hypothesized that destabilization of plasma and acrosomal membranes prior to injection would positively impact the developmental potential of equine zygotes generated by ICSI. This study evaluated effects of the sperm treatment with lysolecithin on plasma and acrosomal membranes and on oocyte activation ability, initially following heterologous ICSI on bovine oocytes and subsequently employing equine oocytes. The effects of the lysolecithin -treatment on the efficiency of conventional and piezo-assisted equine ICSI were evaluated. To do this, the equine sperm were treated with different concentrations of lysolecithin and the sperm plasma membrane, acrosome and DNA integrity were evaluated by flow cytometry. The results showed that a lysolecithin concentration of 0.08 % destabilized the membranes of all sperm and affected DNA integrity within the range described for the species (8-30 %). In addition, the heterologous ICSI assay showed that lysolecithin treatment was detrimental to the sperm's ability to activate the oocyte, therefore, chemical oocyte activation was used after equine ICSI after injection with lysolecithin -treated sperm. This group showed similar developmental rate to the control group with and without exogenous activation. In conclusion, lysolecithin pre-treatment is not necessary when using ICSI to produce equine embryos in vitro. The results from the current study provide additional insight regarding the factors impacting ICSI in horses.

Save the Mekong Delta from drowning.

Policy must address drivers, not just symptoms, of subsidence.

Induction of oxidative stress does not increase the cryotolerance of vitrified embryos.

Short-term treatment of mammalian oocytes with different stressors induces stress tolerance of embryos derived from these oocytes. The aims of this study were to evaluate effects on embryo development when there was treatment of oocyte complexes (COCs) used to derive the embryos with hydrogen peroxide (H2O2).The COCs were not incubated with H2O2: control (0 µM), or were incubated with 25, 50, 75, or 100 µM concentrations of H2O2 for 1 h prior to in vitro fertilization, and presumptive zygotes were cultured until day 7. Blastocysts at day 7 of development derived from H2O2-treated (25 µM treatment concentration) COCs were vitrified. Percentage of embryos undergoing cleavage was not affected by any treatment, while percentage of embryos developing to the blastocyst stage was less when there was treatment of COCs with 100 µM of H2O2. Embryo quality was less when COCs used to derive blastocysts were treated with 50, 75, or 100 µM concentrations of H2O2. There were lesser relative abundances of some mRNA transcripts of interest in blastocysts when there was treatment of COCs with H2O2. After vitrification, there were no differences in embryo re-expansion and hatching rates compared with fresh and vitrified blastocysts of the control group and those derived from COCs treated with 25 µM H2O2. In conclusion, treatment of COCs used to derive blastocysts with H2O2 does not induce stress tolerance in vitrified embryos of cattle; however, the viability of these blastocysts is similar to those of the control group.

Response to Comments on "Designing river flows to improve food security futures in the Lower Mekong Basin".

Sabo et al presented an empirically derived algorithm defining the socioecological response of the Tonle Sap Dai fishery in the Cambodian Mekong to basin-scale variation in hydrologic flow regime. Williams suggests that the analysis leading to the algorithm is flawed because of the large distance between the gauge used to measure water levels (hydrology) and the site of harvest for the fishery. Halls and Moyle argue that Sabo et al's findings are well known and contend that the algorithm is not a comprehensive assessment of sustainability. We argue that Williams' critique stems from a misunderstanding about our analysis; further clarification of the analysis is provided. We regret not citing more of the work indicated by Halls and Moyle, yet we note that our empirical analysis provides additional new insights into Mekong flow-fishery relationships.

An exudate detection method for diagnosis risk of diabetic macular edema in retinal images using feature-based and supervised classification.

The present paper aims at presenting the methodology and first results of a detection system of risk of diabetic macular edema (DME) in fundus images. The system is based on the detection of retinal exudates (Ex), whose presence in the image is clinically used for an early diagnosis of the disease. To do so, the system applies digital image processing algorithms to the retinal image in order to obtain a set of candidate regions to be Ex, which are validated by means of feature extraction and supervised classification techniques. The diagnoses provided by the system on 1058 retinographies of 529 diabetic patients at risk of having DME show that the system can operate at a level of sensitivity comparable to that of ophthalmological specialists: it achieved 0.9000 sensitivity per patient against 0.7733, 0.9133 and 0.9000 of several specialists, where the false negatives were mild clinical cases of the disease. In addition, the level of specificity reached by the system was 0.6939, high enough to screen about 70% of the patients with no evidence of DME. These values show that the system fulfils the requirements for its possible integration into a complete diabetic retinopathy pre-screening tool for the automated management of patients within a screening programme. Graphical Abstract Diagnosis system of risk of diabetic macular edema (DME) based on exudate (Ex) detection in fundus images.

Gradient sperm selection for reproductive techniques in cattle: Is Isolate a suitable replacement for Percoll?

In assisted reproductive techniques, it is essential to perform a sperm selection to obtain spermatozoa with high motility and membrane integrity for in vitro fertilisation (IVF) and high-DNA integrity for intracytoplasmic sperm injection (ICSI). In this study, we evaluated whether Isolate® was a suitable substitute for Percoll® for assisted reproductive techniques. Commercial cryopreserved bovine semen was used after selection in both gradients, and plasma and acrosome membrane integrity, reactive oxygen species (ROS) levels, DNA integrity and mitochondrial membrane potential (ΔΨm) were assessed by flow cytometry. Motility parameters were also evaluated by CASA system. A similar percentage of spermatozoa with intact plasma membrane, acrosome integrity and high ΔΨm was observed in both sperm selection methods, but only Percoll® showed higher percentage of spermatozoa with intact plasma and acrosome membrane compared to the post-thawing group. No differences were observed in the motility, ROS, DNA fragmentation and on the in vitro embryo production in all experimental groups. In conclusion, the selection of bovine spermatozoa with Isolate® generates spermatozoa with similar quality parameters and embryonic development compared to Percoll® providing a suitable alternative sperm selection method for assisted reproductive techniques in this species.

Designing river flows to improve food security futures in the Lower Mekong Basin.

Rivers provide unrivaled opportunity for clean energy via hydropower, but little is known about the potential impact of dam-building on the food security these rivers provide. In tropical rivers, rainfall drives a periodic flood pulse fueling fish production and delivering nutrition to more than 150 million people worldwide. Hydropower will modulate this flood pulse, thereby threatening food security. We identified variance components of the Mekong River flood pulse that predict yield in one of the largest freshwater fisheries in the world. We used these variance components to design an algorithm for a managed hydrograph to explore future yields. This algorithm mimics attributes of discharge variance that drive fishery yield: prolonged low flows followed by a short flood pulse. Designed flows increased yield by a factor of 3.7 relative to historical hydrology. Managing desired components of discharge variance will lead to greater efficiency in the Lower Mekong Basin food system.

Effect of sperm pretreatment with glutathione and membrane destabilizing agents lysolecithin and Triton X-100, on the efficiency of bovine intracytoplasmic sperm injection.

Intracytoplasmic sperm injection (ICSI) is an assisted reproduction tool with several applications. Its effectiveness in bovines is lower than that in other species, mainly because of difficulties in the decondensation of the sperm nucleus after injection, and the presence of the acrosome and the plasma membrane which remain intact in this procedure. In this study, we assessed the effect of lysolecithin (LL) and Triton X-100 (TX), in combination with glutathione (GSH) as sperm pretreatments prior to ICSI. The GSH-LL and GSH-TX groups showed 0% of spermatozoa with intact membrane (SYBR 14+/PI), in comparison with the control (63.3%) and GSH (65.7%) groups. The proportions of spermatozoa with damaged acrosome membrane in the GSH-LL, GSH-TX, GSH and control groups were 46%, 35.9%, 10.5% and 7.5%, respectively. Sperm chromatin decondensation analysis showed that the groups incubated for 3 hr with GSH presented greater decondensation (p < .05). Although fertilization was improved in all treatment groups evaluated, no differences were observed in the cleavage rate 72 hr after activation in the GSH (73.7%), GSH-LL (80.2%) and GSH-TX (77.8%) groups compared to the control (66.3%), neither in the blastocyst rate on day 8 (24.0%, 26.2%, 27.1% and 28.4% for the control, GSH, GSH-LL and GSH-TX groups, respectively). No differences were also observed in the total number of cells in all groups. In conclusion, although these sperm treatments promoted nuclear decondensation and induced plasma membrane disruption, these effects were not sufficient to improve bovine embryonic development after ICSI.

Effect of short-term exposure of cumulus-oocyte complex to 3-morpholinosydnonimine on in vitro embryo development and gene expression in cattle.

Short-term exposure of gametes to different types of stress might induce stress tolerance in mammalian embryos. The aim of this study was to evaluate the effect of short-term exposure of bovine mature cumulus-oocyte complex (COC) to 3-morpholinosydnonimine (SIN-1) on subsequent in vitro embryo development, embryo quality and relative gene expression. Matured COCs were incubated with SIN-1 (0, 0.1, 1, 10 and 100 µM SIN-1) for 1 hr before in vitro fertilization and zygotes were cultured until Day 7. The cleavage rate at 72 hr did not show any differences among groups. However, the blastocyst rate on Day 7 decreased with all treatments evaluated, with the embryos generated with 10 µM SIN-1 showing the lowest embryo production rate. Embryo quality analysis did not show any differences in total cell number (TCN) or inner cell mass (ICM) among groups. Relative gene expression analysis showed a downregulation of eNOS expression and an upregulation of nNOS expression in all treatments evaluated compared to the control group. Also, a downregulation was observed in some treatments: SOD2 at 0.1 µM; SOD1 at 0.1 and 100 µM; PRDX5 at 0.1, 10 and 100 µM; and NANOG at 10 and 100 µM; and an upregulation of CDX2 expression was observed at 100 µM. The other genes (OCT4, HIF1A, HSPA1A, BCL2A and iNOS) did not show any differences in the relative gene expression. These results suggest that the short-term exposure of mature bovine COCs to SIN-1 does not induce stress tolerance and has no beneficial effect on bovine in vitro embryo production.

Viability and DNA fragmentation of rainbow trout embryos (Oncorhynchus mykiss) obtained from eggs stored at 4 °C.

In vitro storage of salmonid eggs leads to aging of the cells causing a decline in quality and reducing their capacity to develop and produce embryos. The quality of salmonid embryos is assessed by morphologic analyses; however, data on the application of biomarkers to determine the cell viability and DNA integrity of embryos in these species are limited. The aim of this study was to evaluate the effect on embryo development, viability and DNA fragmentation in the embryonic cells of in vitro storage time at 4 °C of rainbow trout (Oncorhynchus mykiss) eggs. The embryos were obtained by IVF from eggs stored for 0 (control), 48, and 96 hours at 4 °C. At 72 hours after fertilization, dechorionated embryos were examined to determine percentages of developed embryos (embryos with normal cell division morphology), viability (LIVE/DEAD sperm viability kit), and DNA integrity (terminal deoxynucleotidyl transferase [TdT] dUTP nick-end labeling assay). The percentage of developing embryos decreased (P < 0.05) with storage time of the eggs (95.10 ± 2.55; 88.14 ± 4.50; 79.99 ± 6.60 for 0, 48, and 96 hours, respectively). Similarly, cell viability decreased (P < 0.05; 96.07 ± 7.15; 80.42 ± 8.55; 77.47 ± 7.88 for 0, 48, and 96 hours, respectively), and an increase (P < 0.05) in DNA fragmentation in the embryos was observed at 96-hour storage. A positive correlation was found between cell DNA fragmentation and storage time (r = 0.8173; P < 0.0001). The results revealed that terminal deoxynucleotidyl transferase [TdT] dUTP nick-end labeling assay technique is reliable mean to assess the state of the DNA in salmonid embryos and that in vitro eggs storage for 96h reduces embryo development and cell DNA integrity. DNA integrity evaluation constitutes a biomarker of the quality of the ova and resulting embryos so as to predict their capacity to produce good-quality embryos in salmonids, particularly under culture conditions.

Methyl-ß-Cyclodextrin Improves Sperm Capacitation Status Assessed by Flow Cytometry Analysis and Zona Pellucida-Binding Ability of Frozen/Thawed Bovine Spermatozoa.

Mammalian sperm undergo a series of biochemical transformations in the female reproductive tract that are collectively known as capacitation. Cyclodextrins added to the sperm culture medium have been described to induce in vitro sperm capacitation, enabling its use in protein-free media. However, the additive capacitating effect of methyl-ß-cyclodextrin (MßCD) in the medium containing bovine serum albumin (BSA) is unknown in the bovine species. In this study, we evaluated the effects of incubating frozen-thawed bovine spermatozoa in a BSA-containing medium supplemented with MßCD on different sperm quality and functional parameters. Sperm viability decreased with the addition of MßCD in a dose-dependent manner (p < 0.05), and DNA damage could be observed but only with the highest concentration of MßCD. However, pre-incubation of spermatozoa in MßCD-supplemented medium improved the capacitation status as assessed by the increase in plasma membrane fluidity, intracellular calcium concentration, induced acrosome reactivity and zona pellucida (ZP)-binding ability (p < 0.05). Thus, we conclude that MßCD supplementation is able to enhance the capacitation status of frozen-thawed bovine spermatozoa cultured in capacitation medium containing BSA and could result in a valid strategy for its application on artificial reproductive technologies such as in vitro fertilization or intracytoplasmic sperm injection.

Activation treatment of recipient oocytes affects the subsequent development and ploidy of bovine parthenogenetic and somatic cell nuclear transfer (SCNT) embryos.

We evaluated the potential effect of anisomycin, an antibiotic produced by Streptomyces griseolus, on the parthenogenetic activation of bovine oocytes and reconstructed somatic-cell nuclear transfer (SCNT) embryos. A higher cleavage and blastocyst rate were achieved with anisomycin (70.3% and 27.8%) and 6-dimethylaminopurine (DMAP) (73.3% and 30.2%), relative to oocytes parthenogenetically activated with cycloheximide (CHX) (54.1% and 20.2%). In reconstructed SCNT embryos, a greater proportion of embryos reached the blastocyst stage after anisomycin (32.2%) compared to DMAP (22.3%) and CHX (23.5%) treatment. Furthermore, the quality of embryos-assessed by the total number of cells and the inner cell mass-to-total-cell ratio-was higher with anisomycin (166.2 ± 6.9 and 26.9 ± 1.9) compared to DMAP (135.0 ± 8.7 and 39.4 ± 3.5) and CHX (149.1 ± 8.4 and 36.3 ± 2.5), while a lower percentage of chromosomal abnormalities was observed with anisomycin compared to DMAP and CHX treatments, both in parthenotes (though not significant) and in SCNT embryos (P < 0.05). Therefore, anisomycin can enhance the in vitro developmental potential in parthenotes and reconstructed SCNT embryos, specifically improving the quality of SCNT embryos and decreasing the abnormal ploidy of parthenotes and SCNT embryos compared to the traditional protocols of chemical activation with DMAP and CHX. These results may have important implications for the success of reproductive technologies, including SCNT and intracytoplasmic sperm injection, in different mammalian species.

N-acetyl-L-cysteine pre-treatment protects cryopreserved bovine spermatozoa from reactive oxygen species without compromising the in vitro developmental potential of intracytoplasmic sperm injection embryos.

Excess of reactive oxygen species (ROS) on in vitro embryo production systems negatively affects the quality and developmental potential of embryos, as result of a decreased sperm quality and increased DNA fragmentation. This issue is of major importance in assisted fertilisation procedures such as intracytoplasmic sperm injection (ICSI), because this technique does not allow the natural selection of competent spermatozoa, and therefore, DNA-damaged spermatozoa might be used to fertilise an egg. The aim of this study was to investigate a new strategy to prevent the potential deleterious effect of ROS on cryopreserved bovine spermatozoa. We evaluated the effect of a sperm pre-treatment with different concentrations of N-acetyl-L-cysteine (NAC) on ROS production, viability and DNA fragmentation and assessed the effect of this treatment on the in vitro developmental potential and quality of embryos generated by ICSI. The results show a strong scavenging effect of 1 and 10 mm NAC after exposure of spermatozoa to a ROS inducer, without compromising the viability and DNA integrity. Importantly, in vitro developmental potential and quality of embryos generated by ICSI with spermatozoa treated with NAC were not affected, confirming the feasibility of using this treatment before an ICSI cycle.

Supplementation of IVF medium with melatonin: effect on sperm functionality and in vitro produced bovine embryos.

Gamete co-incubation generates high free radical levels surrounding growing zygotes which may impair subsequent embryo viability. Melatonin eliminates a wide variety of free radicals; hence, we tried to improve in vitro embryo production by adding melatonin to in vitro fertilisation (IVF) media in high (Exp. 1) and low concentrations (Exp. 2), and we evaluated its effect on bull sperm function during IVF co-incubation time (Exp. 3). In Experiment 1, we supplemented IVF media culture with 0.01, 0.1 and 1 mmol of melatonin, along with a no melatonin control group. In Experiment 2, melatonin levels were reduced to 10, 100 and 1000 nmol, with a no melatonin control group. In Experiment 3, spermatozoa were incubated in IVF media with melatonin (as Exp. 2) and functional parameters were analysed at 0, 4 and 18 h. In Experiment 1, only 1 mmol melatonin showed lesser blastocyst rates than control (C: 23.2 ± 6.7% versus 1 mmol: 2.0 ± 1.7%). In Experiment 2, no statistical differences were found in cleavage percentage, blastocyst percentage and total cell count for any melatonin treatment. In Experiment 3, sperm samples with 1000 nmol melatonin had a significantly higher wobbler (WOB) coefficient, a lower percentage of intact acrosomes, a lower percentage of viable spermatozoa with ROS, greater DNA fragmentation and higher DNA oxidation than controls. Total fluorescence intensity for ROS at 10 nmol melatonin was significantly greater than controls (P < 0.05). IVF media with 1 mmol melatonin is deleterious for embryo development, and in lower concentrations, it modulated sperm functionality, but had no effects on embryo production.

Effect of sperm pretreatment with sodium hydroxide and dithiothreitol on the efficiency of bovine intracytoplasmic sperm injection.

The efficiency of intracytoplasmic sperm injection (ICSI) in bovines is lower than in other species due, in part, to a lack of optimal conditions for its implementation; this has hindered the achievement of high rates of embryonic development and the birth of live offspring. The aim of the present study was to evaluate the effects of pretreatment of bovine spermatozoa with NaOH and dithiothreitol (DTT) on the viability, plasma membrane integrity, DNA fragmentation and in vitro developmental potential of embryos generated by ICSI. Following pretreatment of spermatozoa with 5 mM DTT for 20 min and a low concentration of NaOH (1 mM for 60 min), there were fewer live and acrosome reacted spermatozoa (44% and 34%, respectively) than in the control group without treatment (82%). Spermatozoa subjected to higher alkali concentrations (10-50 mM) were mostly dead and reacted. However, pronuclear formation, cleavage, blastocyst rate and embryo quality did not differ between these pretreatment groups and the untreated control group. In conclusion, we have described, for the first time, the effects of NaOH treatment on bovine spermatozoa and subsequent in vitro embryonic development after ICSI, and have demonstrated that pretreatment of bovine spermatozoa with NaOH or DTT is not necessary for an appropriate in vitro embryo development in this species.

A Study of the Sugarcane Yellow Leaf Disease in Argentina.

Yellow leaf disease, caused by Sugarcane yellow leaf virus (SCYLV), is widespread around the world but very little information is available on this viral disease in Argentina. Therefore, the aims of the study were to assess the presence of SCYLV, analyze its distribution in the main sugarcane production areas of Argentina, characterize the virus, and determine histological alterations caused by its presence. For this purpose, 148 sugarcane samples with and without symptoms were collected in 2011 and 2012 from the province of Tucumán. One additional sample was collected in Salta, a different geographical, agroecological, and producing region. Results showed that SCYLV is widely distributed in commercial varieties of sugarcane throughout Tucumán in both symptomatic and asymptomatic leaves. A low but statistically significant positive correlation with virus detection and disease symptoms was found. BRA-PER was the only genotype detected by reverse-transcription polymerase chain reaction and sequence analysis of the SCYLV capsid protein gene. SCYLV-positive samples showed high starch levels in bundle sheath cells, whereas the asymptomatic ones, probably in an early stage of infection, were found to contain more chloroplasts. Symptomatic noninfected samples presented crystal formation probably associated with phytoplasma infection.

Influence of xenobiotic contaminants on landfill soil microbial activity and diversity.

Landfills are often the final recipient of a range of environmentally important contaminants such as hydrocarbons, polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs). In this study the influence of these contaminants on microbial activity and diversity was assessed in a municipal solid waste (MSW) landfill placed in Torrejón de Ardoz (Madrid, Spain). Soil samples were collected from four selected areas (T2, T2B, T8 and T9) in which the amount of total hydrocarbons, PAHs and PCBs were measured. Soil biomass, substrate induced respiration (SIR) and physiological profiles of soil samples were also determined and used as indicators of total microbial activity. Highest concentration of total hydrocarbons was detected in T2 and T9 samples, with both PCBs and benzopyrene being detected in T9 sample. Results corresponding to microbial estimation (viable bacteria and fungi, and SIR) and microbiological enzyme activities showed that highest values corresponded to areas with the lowest concentration of hydrocarbons (T2B and T8). It is noticeable that in such areas was detected the lowest concentration of the pollutants PAHs and PCBs. A negative significant correlation between soil hydrocarbons concentration and SIR, total bacteria and fungi counts and most of the enzyme activities determined was established. DGGE analysis was also carried out to determine the microbial communities' structure in the soil samples, establishing different profiles of Bacteria and Archaea communities in each analysed area. Through the statistical analysis a significant negative correlation was only found for Bacteria domain when Shannon index and hydrocarbon concentration were correlated. In addition, a bacterial 16S rRNA gene based clone library was prepared from each soil. From the clones analysed in the samples, the majority corresponded to Proteobacteria, followed by Acidobacteria and Actinobacteria. It is important to remark that the most polluted sample (T9) showed the lowest microbial diversity only formed by six phyla being Proteobacteria and Acidobacteria the most representative.

First Report of Anthracnose on Fruits of Duchesnea indica Caused by Colletotrichum acutatum in Northwestern Argentina.

Duchesnea indica (Andrews) Focke, a cosmopolitan wild species related to the cultivated strawberry that is widely distributed in northwestern Argentina, grows in close proximity to strawberry crops and has proven to be almost immune to Colletotrichum spp. isolated from diseased strawberry plants (1), hence it has never been considered a phytopathological risk. During a field survey of "La Heladera" (27°01'45″S, 65°39'20″W), Tafí del Valle (Tucumán, Argentina) from November 2009 to November 2010, a genotype of D. indica showing fruits with dark brown, necrotic, irregular, circular lesions of 5 to 20 mm in diameter were collected. Setose acervuli were observed on the center of the fruit lesions. Pathogens were obtained from 10 diseased fruit collected at random, and four fungal isolates were isolated per fruit on potato dextrose agar (PDA). To reduce the number of samples for evaluation, two isolates per fruit that were exhibiting stable but distinctive morphological features were chosen to continue the studies. Isolates were characterized by morphological, molecular, and phytopathological criteria. After 10 days of incubation on PDA medium at 28°C with continuous white light, colonies exhibited a gray, aerial mycelium, whereas the reverse of the colony is a pale maroon with a radial, pale salmon color. Masses of salmon-colored conidia formed in the center of the colonies. Conidia were hyaline, one celled, fusiform, tapered to a point at both ends, and measured 14.8 to 17.3 × 4.5 to 7.4 µm (n = 100). Setae were scarce and sclerotia were absent. All morphological characteristics that were observed indicated that the isolates were C. acutatum (3). To fulfill Koch's postulates and verify the pathogenicity on commercial varieties of strawberry, six healthy plants of D. indica and Fragaria × ananassa cv. Camarosa with mature fruits were used to test each isolate. Four plants were spray inoculated with conidial suspensions of the virulent isolates (1.5 × 106 conidia/ml) and two with sterile distilled water as controls. Both treatments were maintained under white light (2,000 lux, 12 h per day) at 28°C and 70% relative humidity. Nine days after the inoculation, dark brown lesions and salmon-colored masses of conidia were observed only in inoculated fruits of both genotypes. The fungus isolated from diseased fruits and the conidia that were produced were identical to the isolates used to inoculate the plants. To confirm pathogen identity, PCR amplification with the species-specific pair of primer CaInt2/ITS4 (4) were carried out using fungal total DNA from the original isolates and isolates obtained from inoculated fruits. An amplification product of approximately 490 bp, which is specific for C. acutatum, was observed in all DNA samples (4). Although C. acutatum has already been reported in Fragaria × ananassa in Argentina (2), to our knowledge, this is the first report of C. acutaum causing anthracnose in D. indica species. This result is relevant since this species grows close to strawberry fields and can be an alternative host and potential vector of the anthracnose disease agent. References: (1) M. E. Arias. Frutillas Silvestres y Especies Relacionadas con la Cultivada. EDUNT, Argentina, 2007. (2) C. J. Ramallo et al. Plant Dis. 84:706. 2000. (3) B. J. Smith and L. L. Black. Plant Dis. 74:69, 1990. (4) S. Sreenivasaprasad et al. Plant Pathol. 45:650, 1996.

Effect of different sequential and two-step culture systems on the development, quality, and RNA expression profile of bovine blastocysts produced in vitro.

In the present study, we examined the effect of two-step and sequential culture systems on the development, quality, and gene expression profile of bovine embryos generated by in vitro fertilization. Presumptive zygotes were randomly allocated to four culture treatments: (1) KSOM + 0.4% BSA for 3 days, and then KSOM + 5% FBS to day 7 (K-K/FBS); (2) KSOM + 0.1% BSA for 3 days, and then SOF + 5% FBS to day 7 (K-S/FBS); (3) KSOM + 0.1% BSA for 3 days, and then SOF + 0.8% BSA to day 7 (K-S/BSA); and (4) KSOM + 0.4% BSA for 3 days, and then KSOM + 0.8% BSA to day 7 (K-K/BSA). Culture medium had no effect on cleavage rate. However, a significant difference (P < 0.01) was observed with the two-step culture systems, yielding higher rate of blastocysts (37 and 32% for K-K/FBS and K-K/BSA, respectively) compared to sequential culture systems (26 and 28% for K-S/FBS and K-S/BSA, respectively). Embryos cultured in sequential K-S/FBS developed slowly, had a lower hatching rate, fewer cells, and a higher apoptosis rate compared to other treatments. Gene expression analysis showed alterations of DNMT1, OCT-4, and SOD2 in embryos cultured in sequential K-S/FBS and SOD1 in embryos cultured in two-step K-K/BSA. In conclusion, in vitro culture systems may have an impact not just in the developmental potential and quality of the generated embryos but also in the gene expression profile, which suggests that changes in the culture medium composition can modulate global gene expression.

Avances en inmunidad gestacional / Advances in gestational immunity

El inicio y establecimiento de la gestación en los mamíferos dependen de la adaptación del sistema inmunológico de la madre para tolerar un feto semi-alogénico. La gestación en sí misma constituye un acontecimiento de equilibrio inmunológico, ya que mientras el sistema inmune mantiene la competencia para la defensa contra antígenos foráneos, mecanismos de tolerancia local y periférica previenen una respuesta inapropiada contra alo-antígenos fetales de origen paterno lo que pudiera provocar el rechazo del feto. La interacción materno-fetal es extremadamente compleja y es difícil determinar todos los componentes del sistema inmune involucrados. Hasta ahora se ha demostrado la participación activa de las células T y sus productos, las citoquinas y también se ha involucrado a las moléculas del complejo mayor de histocompatibilidad, los antígenos paternos y algunos inmunomoduladores como progesterona, indoleamina 2,3-dioxigeneasa y glicodelina, entre otros. Todos estos elementos parecen confluir para producir un gran cambio sistémico en el sistema inmune materno, promoviendo por una parte la tolerancia materno-fetal, crucial para finalmente permitir una gestación exitosa y, por otro lado, manteniendo una activa vigilancia inmune contra las infecciones que pondrían en riesgo la gestación y sobrevivencia de diversas especies. Se revisó la literatura más reciente acerca de los diferentes componentes del sistema inmune que han demostrado ser clave en el inicio y mantención de la gestación en mamíferos.

The initiation and establishment of pregnancy in mammals depends on the adaptation from maternal immune system to tolerate a semi-allogeneic fetus. Pregnancy itself constitutes an event of immune balance because, while the immune system maintains the capacity for defense against foreign antigens, mechanisms of local and peripheral tolerance may prevent an inappropriate response against fetal alloantigens of paternal origin which could lead to rejection of the fetus. The maternal-fetal immune interaction is extremely complex and it has therefore been difficult to identify all the immune components involved. So far, it is known that the active participation of T cells and their products, cytokines, and has also involved molecules from the major histocompatibility complex, other paternal antigens and some immunomodulators molecules such as progesterone, glycodelin and indoleamine 2,3-dioxigenase among others. All these elements seem to converge to produce a major systemic change in the maternal immune system, promoting on one hand the maternal-fetal tolerance, crucial to allow a successful pregnancy and on the other hand, maintaining an active immune surveillance against infections that might endanger pregnancy and survival of diverses species. A review of recent literature about the different components of the immune system that have proven key in the beginning and maintenance of pregnancy in mammals.