Reference genome of Townsend's big-eared bat, Corynorhinus townsendii.

Townsend's big-eared bat, Corynorhinus townsendii, is a cave- and mine-roosting species found largely in western North America. Considered a species of conservation concern throughout much of its range, protection efforts would greatly benefit from understanding patterns of population structure, genetic diversity, and local adaptation. To facilitate such research, we present the first de novo genome assembly of C. townsendii as part of the California Conservation Genomics Project (CCGP). Pacific Biosciences HiFi long reads and Omni-C chromatin-proximity sequencing technologies were used to produce a de novo genome assembly, consistent with the standard CCGP reference genome protocol. This assembly comprises 391 scaffolds spanning 2.1 Gb, represented by a scaffold N50 of 174.6 Mb, a contig N50 of 23.4 Mb, and a benchmarking universal single-copy ortholog (BUSCO) completeness score of 96.6%. This high-quality genome will be a key tool for informed conservation and management of this vulnerable species in California and across its range.

Immunological uterine response to pig embryos before and during implantation.

The establishment of a successful pregnancy can only occur through a concerted functioning of the entire female reproductive system, allowing for fertilization, subsequent embryo development and implantation of the conceptus. In this context, the uterine immunological responses responsible for rejection or tolerance of the conceptus are of critical importance. The aim of the present review is to summarize our current knowledge about those cellular and molecular immunological events occurring at the uterine level during pre-implantation and implantation stages of pregnancy in the pig. Advancing our understanding of the immune mechanisms involved in the success or failure of pregnancy will provide cues to develop novel strategies augmenting endometrial receptivity, finally increasing the efficiency of assisted reproductive technologies in pigs.

Seminal plasma affects sperm sex sorting in boars.

Two experiments were conducted in boar semen samples to evaluate how both holding time (24h) and the presence of seminal plasma (SP) before sorting affect sperm sortability and the ability of sex-sorted spermatozoa to tolerate liquid storage. Whole ejaculate samples were divided into three aliquots immediately after collection: one was diluted (1:1, v/v) in Beltsville thawing solution (BTS; 50% SP); the SP of the other two aliquots was removed and the sperm pellets were diluted with BTS + 10% of their own SP (10% SP) or BTS alone (0% SP). The three aliquots of each ejaculate were divided into two portions, one that was processed immediately for sorting and a second that was sorted after 24h storage at 15-17°C. In the first experiment, the ability to exhibit well-defined X- and Y-chromosome-bearing sperm peaks (split) in the cytometry histogram and the subsequent sorting efficiency were assessed (20 ejaculates). In contrast with holding time, the SP proportion influenced the parameters examined, as evidenced by the higher number of ejaculates exhibiting split and better sorting efficiency (P<0.05) in semen samples with 0-10% SP compared with those with 50% SP. In a second experiment, the quality (viability, total and progressive motility) and functionality (plasma membrane fluidity and intracellular generation of reactive oxygen species) of sex-sorted spermatozoa were evaluated after 0, 72 and 120h storage at 15-17°C (10 ejaculates). Holding time and SP proportion did not influence the quality or functionality of stored sex-sorted spermatozoa. In conclusion, a holding time as long as 24h before sorting did not negatively affect sex sorting efficiency or the ability of sorted boar spermatozoa to tolerate long-term liquid storage. A high proportion of SP (50%) in the semen samples before sorting reduced the number of ejaculates to be sorted and negatively influenced the sorting efficiency, but did not affect the ability of sex-sorted spermatozoa to tolerate liquid storage.

Boar sperm cryosurvival is better after exposure to seminal plasma from selected fractions than to those from entire ejaculate.

Boar bulk ejaculates are now being collected instead of usual sperm-rich fractions (SRF) for artificial insemination purpose. The present study evaluated the influence of holding boar sperm samples before freezing surrounded in their own seminal plasma (SP), from either fractions/portions or the entire ejaculate, on post-thawing sperm quality and functionality. Ejaculates collected as bulk (BE) or as separate (first 10 mL of SRF [P1] and rest of SRF [P2]) from 10 boars were held 24h at 15-17°C and then frozen. Some bulk ejaculate samples were frozen immediately after collections as Control. In addition, epididymal sperm samples from the same 10 boars were collected post-mortem and extended in SP from P1 (EP1), P2 (EP2) and post SRF (EP3), and also held 24h before freezing for a better understanding of the influence of SP on boar sperm cryopreservation. The sperm quality (motility, evaluated by CASA, and viability, evaluated by flow cytometry) and functionality (flow cytometry assessment of plasma membrane fluidity, mitochondrial membrane potential and intracellular generation of reactive oxygen species [ROS] in viable sperm) were evaluated at 30, 150 and 300 min post-thaw. Post-thawing sperm quality and functionality of P1 and P2 were similar but higher (p < 0.01) than BE samples. Control samples showed higher (p < 0.01) post-thaw sperm quality and functionality than BE samples. Post-thawing sperm quality and functionality of EP1 and EP2 were similar but higher (p < 0.05) than EP3. These results showed that boar sperm from BE are more cryosensitive than those from the SRF, particularly when held 24h before freezing, which would be attributable to the cryonegative effects exerted by the SP from post SRF.

Egg yolk and glycerol requirements for freezing boar spermatozoa treated with methyl ß-cyclodextrin or cholesterol-loaded cyclodextrin.

Egg yolk (EY) and glycerol are common constituents of extenders used for sperm cryopreservation. It has been demonstrated that using cholesterol-loaded cyclodextrins (CLC) improves sperm cryosurvival in several species. However, standard freezing extenders might not be the most appropriate for CLC-treated sperm. This study evaluated the EY and glycerol requirements for freezing CLC-treated boar spermatozoa. Semen samples from 34 ejaculates coming from 4 boars were used. Each ejaculate was split into three aliquots: one was used untreated (control), and the other two were treated with 1 mg of CLC or methyl-ß-cyclodextrin/120 × 10(6) sperm for 15 min at 22 C prior to cryopreservation. Our results indicated that reducing the concentration of EY was detrimental for sperm viability after thawing (31.57 ± 2 vs. 19.89% ± 2 for 20 and 10% EY, respectively; P <0.05), even in semen treated with CLC. On the other hand, it was observed that the traditional concentration of glycerol (3%) was not the appropriate for freezing CLC-treated sperm (61.10 ± 3 vs. 47.87% ± 3 viable sperm for control and CLC-treated sperm, respectively; P <0.05). Thus, CLC-treated sperm showed a higher tolerance to high glycerol concentrations (5%) in terms of sperm viability (59.19% ± 3) than non-treated sperm (45.58% ± 3; P<0.05). Therefore, it could be necessary to modify the freezing extenders for CLC-treated sperm. Nevertheless, additional studies will be needed to evaluate alternative cryoprotectants and to determine the effect of high glycerol concentrations on sperm functionality.

Semen proteome and transcriptome of the endangered black-footed ferret (Mustela nigripes) show association with the environment and fertility outcome.

The ex situ population of the endangered black-footed ferret (Mustela nigripes) has been experiencing declines in reproductive success over the past 30 years of human-managed care. A potential cause may be environmental-dependent inbreeding depression with diet being one of the contributing factors since ferrets are not fed their natural diet of prairie dogs. Here, we generated and analyzed semen proteome and transcriptome data from both wild and ex situ ferrets maintained on various diets. We identified 1757 proteins across all samples, with 149 proteins unique to the semen of wild ferrets and forming a ribosomal predicted protein-protein interaction cluster. Wild ferrets also differed from ex situ ferrets in their transcriptomic profile, showing enrichment in ribosomal RNA processing and potassium ion transport. Successful fertility outcomes documented for ex situ ferrets showed the strongest association with the semen transcriptome, with enrichment in genes involved in translation initiation and focal adhesion. Fertility also synergized with the effect of diet on differentially expressed transcriptomes, mainly affecting genes enriched in mitochondrial function. Our data and functional networks are important for understanding the causes and mechanisms of declining fertility in the ex situ ferret population and can be used as a resource for future conservation efforts.

Season of ejaculate collection influences the freezability of boar spermatozoa.

The aim of this retrospective study was to evaluate whether the season of ejaculate collection influences the freezability of porcine sperm. A total of 434 ejaculates were collected from boars of six different breeds over three years (2008-2011) and throughout the four seasons of the year identified in the northern hemisphere (winter, spring, summer and autumn). The ejaculates were cryopreserved using a standard 0.5 mL straw freezing protocol. Sperm quality was assessed before (fresh semen samples kept 24h at 17°C) and after freezing and thawing (at 30 and 150 min post-thawing in semen samples kept in a water bath at 37 °C), according to the percentages of total motility, as assessed by the CASA system, and viability, as assessed by flow cytometry after staining with SYBR-14, PI and PE-PNA. The data, in percentages, on sperm motility and viability after freezing and thawing were obtained at each evaluation time (recovered) and were normalized to the values before freezing (normalized). The season of ejaculate collection influenced (P<0.01) sperm quality before freezing and after thawing (recovered and normalized), irrespective of the breed of boar. Sperm quality was lower in summer, both in terms of motility and viability, and in autumn, in terms of motility, than in winter and spring. Seasonality in the normalized data indicates that the season of ejaculate collection influences sperm freezability, regardless of the season's influence on sperm quality before freezing. Consequently, the spermatozoa from ejaculates collected during summer and, to a lesser extent, also in autumn, are more sensitive to cryopreservation than those from ejaculates collected during winter and spring.

The Use of a Brief Synchronization Treatment after Weaning, Combined with Superovulation, Has Moderate Effects on the Gene Expression of Surviving Pig Blastocysts.

The combination of estrus synchronization and superovulation (SS) treatments causes alterations in ovarian and endometrial gene expression patterns, resulting in abnormal follicle and oocyte growth, fertilization, and embryo development. However, the impact of combined SS treatments on the transcriptome of the surviving embryos remains unidentified. In this study, we examined gene expression changes in day 6 blastocysts that survived a brief regimen of synchronization treatment combined with superovulation. The sows were included in one of three groups: SS7 group (n = 6), sows were administered Altrenogest (ALT) 7 days from the day of weaning and superovulated with eCG 24 h after the end of ALT treatment and hCG at the onset of estrus; SO group (n = 6), ALT nontreated sows were superovulated with eCG 24 h postweaning and hCG at the onset of estrus; control group (n = 6), weaned sows displaying natural estrus. Six days after insemination, the sows underwent a surgical intervention for embryo collection. Transcriptome analysis was performed on blastocyst-stage embryos with good morphology. Differentially expressed genes (DEGs) between groups were detected using one-way ANOVA with an un-adjusted p-value < 0.05 and a fold change 1.5. The effect of SO treatment on the number of altered pathways and DEGs within each pathway was minimal. Only four pathways were disrupted comprising only a total of four altered transcripts, which were not related to reproductive functions or embryonic development. On the other hand, the surviving blastocysts subjected to SS7 treatments exhibited moderate gene expression changes in terms of DEGs and fold changes, with seven pathways disrupted containing a total of 10 transcripts affected. In this case, the up-regulation of certain pathways, such as the metabolic pathway, with two up-regulated genes associated with reproductive functions, namely RDH10 and SPTLC2, may suggest suboptimal embryo quality, while the down-regulation of others, such as the glutathione metabolism pathway, with down-regulated genes related to cellular detoxification of reactive oxygen species, namely GSTK1 and GSTO1, could depress the embryos' response to oxidative stress, thereby impairing subsequent embryo development. The gene expression changes observed in the present study in SS7 embryos, along with previous reports indicating SS7 can negatively affect fertilization, embryo production, and reproductive tract gene expression, make its use in embryo transfer programs unrecommendable.

Exogenous Melatonin in the Culture Medium Does Not Affect the Development of In Vivo-Derived Pig Embryos but Substantially Improves the Quality of In Vitro-Produced Embryos.

Cloned and transgenic pigs are relevant human disease models and serve as potential donors for regenerative medicine and xenotransplantation. These technologies demand oocytes and embryos of good quality. However, the current protocols for in vitro production (IVP) of pig embryos give reduced blastocyst efficiency and embryo quality compared to in vivo controls. This is likely due to culture conditions jeopardizing embryonic homeostasis including the effect of reactive oxygen species (ROS) influence. In this study, the antioxidant melatonin (1 nM) in the maturation medium, fertilization medium, or both media was ineffective in enhancing fertilization or embryonic development parameters of in vitro fertilized oocytes. Supplementation of melatonin in the fertilization medium also had no effect on sperm function. In contrast, the addition of melatonin to the embryo culture medium accelerated the timing of embryonic development and increased the percentages of cleaved embryos and presumed zygotes that developed to the blastocyst stage. Furthermore, it increased the number of inner mass cells and the inner mass cell/total cell number ratio per blastocyst while increasing intracellular glutathione and reducing ROS and DNA damage levels in embryos. Contrarily, the addition of melatonin to the embryo culture medium had no evident effect on in vivo-derived embryos, including the developmental capacity and the quality of in vivo-derived 4-cell embryos or the percentage of genome-edited in vivo-derived zygotes achieving the blastocyst stage. In conclusion, exogenous melatonin in the embryo culture medium enhances the development and quality of in vitro-derived embryos but not in in vivo-derived embryos. Exogenous melatonin is thus recommended during embryo culture of oocytes matured and fertilized in vitro for improving porcine IVP efficiency.

Boar semen can tolerate rapid cooling rates prior to freezing.

Two experiments were performed in the present study that demonstrated that boar spermatozoa are capable of surviving rapid cooling rates within a range of 15-5 °C before freezing. Boar ejaculates diluted in Beltsville thawing solution (BTS) (1:1, v/v) were held at 17-20 °C and shipped over a 24-h time period from two AI centres to a cryobiology laboratory, where they were pooled (Experiment 1) or cryopreserved individually (Experiment 2) using a standard 0.5-mL straw freezing protocol. The effects of cooling before freezing were assessed after thawing through the objective evaluation of sperm motility and flow cytometric analysis of membrane integrity, acrosomal status, changes in membrane lipid architecture monitored by merocyanine and annexin V binding and intracellular production of reactive oxygen species. In Experiment 1 (six replicates), two semen pools (five ejaculates per pool) were cooled from 15 to 5 °C at rates of 0.08, 0.13, 0.40 and 1.50 °C min(-1). These cooling rates did not result in any significant differences (P>0.05) in any of the post-thaw sperm assessments, even in thawed samples incubated under capacitation conditions. In Experiment 2, three individual ejaculates from 16 boars were slowly (0.08 °C min(-1)) or rapidly (1.5 °C min(-1)) cooled before freezing. A consistent interboar variability (P<0.01) was detected, which was independent of the cooling rate used. Cooling rate only significantly influenced (P<0.05) sperm assessments in four of 16 boars, which exhibited slightly higher percentages of motile cells and intact plasma and acrosomal membranes in the samples that had been cooled slowly. These findings demonstrate that boar spermatozoa undergoing cryopreservation can withstand rapid cooling rates before freezing.

Pervasive duplication of tumor suppressors in Afrotherians during the evolution of large bodies and reduced cancer risk.

The risk of developing cancer is correlated with body size and lifespan within species. Between species, however, there is no correlation between cancer and either body size or lifespan, indicating that large, long-lived species have evolved enhanced cancer protection mechanisms. Elephants and their relatives (Proboscideans) are a particularly interesting lineage for the exploration of mechanisms underlying the evolution of augmented cancer resistance because they evolved large bodies recently within a clade of smaller-bodied species (Afrotherians). Here, we explore the contribution of gene duplication to body size and cancer risk in Afrotherians. Unexpectedly, we found that tumor suppressor duplication was pervasive in Afrotherian genomes, rather than restricted to Proboscideans. Proboscideans, however, have duplicates in unique pathways that may underlie some aspects of their remarkable anti-cancer cell biology. These data suggest that duplication of tumor suppressor genes facilitated the evolution of increased body size by compensating for decreasing intrinsic cancer risk.

From the gigantic blue whale to the minuscule bumblebee bat, animals come in all shapes and sizes. Any species can develop cancer, but some are more at risk than others. In theory, if every cell has the same probability of becoming cancerous, then bigger animals should get cancer more often since they have more cells than smaller ones. Amongst the same species, this relationship is true: taller people and bigger dogs have a greater cancer risk than their smaller counterparts. Yet this correlation does not hold when comparing between species: remarkably large creatures, like elephants and whales, are not more likely to have cancer than any other animal. But how have these gigantic animals evolved to be at lower risk for the disease? To investigate, Vazquez and Lynch compared the cancer risk and the genetic information of a diverse group of closely related animals with different body sizes. This included elephants, woolly mammoths and mastodons as well as their small relatives, the manatees, armadillos, and marmot-sized hyraxes. Examining these species' genomes revealed that, during evolution, elephants had acquired extra copies of 'tumour suppressor genes' which can sense and repair the genetic and cellular damages that turn healthy cells into tumours. This allowed the species to evolve large bodies while lowering their risk of cancer. Further studies could investigate whether other gigantic animals evolved similar ways to shield themselves from cancer; these could also examine precisely how having additional copies of cancer-protecting genes helps reduce cancer risk, potentially paving the way for new approaches to treat or prevent the disease.

Flow Cytometry and Confocal Microscopy for ROS Evaluation in Fish and Human Spermatozoa.

Reactive oxygen species (ROS) could have a negative impact on sperm cellular function and viability. This chapter describes a protocol for oxidative stress evaluation using dichlorofluorescein (DCF) which can specifically reveal intracellular reactive oxygen species. The protocol described here has been used in human and teleost species sperm samples. The method can be used with two approaches: (1) flow cytometry, for quantification of DCF+ cells, or (2) confocal microscopy, for the localization of ROS within the cells.

A Short-Term Altrenogest Treatment Post-weaning Followed by Superovulation Reduces Pregnancy Rates and Embryo Production Efficiency in Multiparous Sows.

Although embryo transfer (ET) is a biotechnology ready for the swine industry, there are factors to be solved, the availability of embryo donors as one. Multiparous sows as donors ought to be considered since weaning is a natural and efficient method for estrus synchronization. In addition, superovulation treatments at weaning are effective in increasing the efficiency of donor embryo production. However, ET programs typically require more donors than those available from a single weaning, imposing grouping several weanings to establish a batch for ET. Since short-term administration of Altrenogest is effective in delaying estrus after weaning without effects on ovulation and embryo development, we investigated how Altrenogest combined with superovulation would affect reproductive parameters and embryo quality and quantity of weaned multiparous donor sows. The sows were administered Altrenogest from the day of weaning for 14 (SS-14 group; N = 26), 7 (SS-7 group; N = 31) and 4 (SS-4 group; N = 32) days. The sows were superovulated with eCG 24 h after the last administration of Altrenogest and with hCG at the onset of estrus. Sows not treated with Altrenogest that were superovulated with eCG 24 h post-weaning and hCG at the onset of estrus (SC group; N = 37) and sows with natural estrus after weaning (C group; N = 34) were used as control groups. The percentage of sows showing estrus within 10 days was not affected by the treatment, but the interval from Altrenogest withdrawal to estrus was longer (P < 0.05) in the SS groups than the interval from weaning to estrus in the controls. SS treatments increased (P < 0.05) the percentage of sows with ovarian cysts and the development of polycystic ovaries. The pregnancy and the fertilization rates, and the overall embryo production efficiency were also negatively affected by the SS treatments (P < 0.05). Interestingly, almost 70% of the structures classified as unfertilized oocytes or degenerated embryos in sows from the SS groups were immature oocytes. In conclusion, although superovulation of weaned sows was highly efficient, short-term administration of Altrenogest in combination with superovulation had negative effects on most of the reproductive parameters assessed, particularly affecting the overall efficiency of pregnancy and embryo production.

Pentoxifylline added to freezing or post-thaw extenders does not improve the survival or in vitro fertilising capacity of boar spermatozoa.

This study evaluated whether pentoxifylline added to freezing and thawing extenders influenced the function of boar spermatozoa. In Experiment 1, pooled ejaculated sperm-rich fractions were frozen in 0.5 ml straws after dilution in extender supplemented with pentoxifylline to a final concentration of 0, 2, 4, 8, 16 or 32 mM. The addition of 4, 8, 16 and 32 mM pentoxifylline to the freezing extender significantly decreased the progressive and total motility of spermatozoa. The percentage of viable spermatozoa with intact acrosomes as well as the penetration rate and the efficiency of fertilisation were significantly lower in pentoxifylline-treated groups compared with the untreated control. In Experiment 2, a pool of three straws with 'good' post-thaw sperm quality parameters and another three straws with 'poor' sperm quality were diluted in extender with 0, 1, 2, 4, 8, 16 or 32 mM pentoxifylline. Post-thaw samples with both 'good' and 'poor' sperm quality with 0, 2, 4, 8 and 16 mM were used to assess IVF parameters. The addition of pentoxifylline to post-thaw extender did not improve the post-thaw motility or viability of spermatozoa compared with the control. The in vitro penetration was higher (P<0.05) than the control for oocytes fertilised with spermatozoa that were thawed and incubated in extender with 4, 8 and 16 mM pentoxifylline. However, no differences were observed in the efficiency of fertilisation. We conclude that pentoxifylline, as a supplement added to the freezing extender, has a deleterious effect and that it does not improve the survival or in vitro fertilising efficiency of frozen-thawed boar spermatozoa when added after thawing.

Probiotics reduce anxiety-related behavior in zebrafish.

There is increasing evidence that gut microbiome could have effects on neurological processes and on behavior. In this study we used the novel tank test (NTT) to analyze zebrafish exploring behavior after four months' supplementation with probiotics with probed antioxidant and anti-inflammatory properties. Results showed that prolonged ingestion of Lactobacillus rhamnosus CECT8361 and Bifidobacterium longum CECT7347 significantly alters the swimming pattern and mean swimming speed in the zebrafish model. After treatment, zebrafish strongly reduced their bottom-dwelling geotactic behavior when placed in a new tank, which could be correlated to a lower state of anxiety.

PSP-I/PSP-II spermadhesin exert a decapacitation effect on highly extended boar spermatozoa.

PSP-I/PSP-II heterodimer is a major protein of boar seminal plasma that is able to preserve, in vitro, the viability, motility and mitochondrial activity of highly-extended boar spermatozoa. However, a relationship between the protective effects of the heterodimer and sperm capacitation is still unclear. The present study investigated the effect of the PSP-I/PSP-II (1.5 mg/mL) on membrane stability, intracellular calcium concentration ([Ca(2+)](I)) and plasma membrane and acrosome integrity of highly extended boar spermatozoa. Boar spermatozoa were diluted to 1 x 10(6) spermatozoa/mL and incubated at 38 degrees C in Phosphate-buffered saline (PBS) for 10, 30, 60, 120 and 300 min or in modified Tris-buffered medium (mTBM) for 10, 20, 30, 60 and 120 min. After each incubation time, the membrane stability (using Merocyanine-540/Yo-Pro-1), elevation of [Ca(2+)](I) (using Fluo-3-AM/PI) and the sperm plasma membrane and acrosome integrity (using SYBR-14/PI/PE-PNA) were evaluated by flow cytometry. As expected, exposure of the spermatozoa to the PSP-I/PSP-II preserved the plasma membrane and acrosome integrity compared to non-exposed spermatozoa in both media PBS and mTBM (p < .01). The evaluation of membrane stability showed no differences in the percentages of viable sperm with instable plasma membrane in the presence of the PSP-I/PSP-II compared to controls irrespective of the dilution media. The evaluation of the [Ca(2+)](I) levels showed that while spermatozoa incubated in mTBM and exposed to PSP-I/PSP-II had lower [Ca(2+)](I) than controls (39.08% vs. 47.97%, respectively; p < .05), no differences were observed in those samples incubated in PBS. However, a temporal evaluation of the samples showed that a similar proportion of live spermatozoa were able to achieve high levels of [Ca(2+)](I) and membrane instability independent of the presence of PSP-I/PSP-II. In conclusion, PSP-I/PSP-II exert a non-permanent decapacitation effect on highly extended boar spermatozoa that is related with a delay in the increase of [Ca(2+)](I) levels.

Seminal Plasma Cytokines Are Predictive of the Outcome of Boar Sperm Preservation.

Background: Boar seminal plasma is rich in cytokines, which could influence the capability of spermatozoa to tolerate preservation. Objectives: To evaluate the involvement of boar seminal plasma cytokines in the changes experienced by boar spermatozoa during their storage, either in liquid or frozen state. Materials and Methods: In two separated experiments, semen samples from healthy and fertile boars were split in two aliquots, one centrifuged twice (1,500 ×g for 10 min) to harvest seminal plasma, whereas the other was either commercially extended (3 × 107 sperm/mL) and liquid-stored at 17°C during 144 h (n = 28, Experiment 1) or frozen-thawed using a standard 0.5 mL protocol (n = 27, Experiment 2). Sixteen cytokines were quantified using Luminex xMAP®. Sperm attributes (CASA-evaluated total and progressive motility; flow cytometry-evaluated sperm viability, production of intracellular H2O2 and O 2 • - and levels of lipid peroxidation in viable spermatozoa) were evaluated either at 0, 72, or 144 h of liquid storage (Experiment 1) or before freezing and at 30- and 150-min post-thawing (Experiment 2). Results: Multiple linear regression models, with Bayesian approach for variable selection, revealed that the anti-inflammatory TGF-ß2, TGF-ß3, IL-1Ra, and IL-4 and the pro-inflammatory IL-8 and IL-18, predicted changes in sperm motility for liquid-stored semen while the anti-inflammatory IFN-γ was included in the models predicting changes in all sperm attributes for cryopreserved semen. Conclusion: Specific boar seminal plasma cytokines would contribute to modulate the structural and metabolic changes shown by spermatozoa during preservation, either in liquid or frozen state.

Long Exposure to a Diet Supplemented with Antioxidant and Anti-Inflammatory Probiotics Improves Sperm Quality and Progeny Survival in the Zebrafish Model.

The aim of the present experiment is to study the effects of oral ingestion of a mixture of two probiotic bacteria on sperm quality and progenies. Three homogeneous groups of juvenile zebrafish were created. Once having reached adulthood (3 months postfertilization; mpf), each group received different feeding regimens: a standard diet (control), a maltodextrin-supplemented diet (vehicle control), or a probiotic-supplemented diet (a mixture (1:1) of Lactobacillus rhamnosus CECT8361 and Bifidobacterium longum CECT7347). The feeding regime lasted 4.5 months. Growth parameters (weight and length) were determined at 3, 5, and 7.5 mpf. Sperm motility was evaluated using computer-assisted sperm analysis at 5 and 7.5 mpf. Progeny survival, hatching rate, and malformation rate were also evaluated. Results showed that probiotic-supplemented diet improved growth parameters compared with the standard diet. The highest percentage of motile spermatozoa was reported in the probiotic-fed group. Concomitantly, the percentage of fast sperm subpopulation was significantly lower in samples derived from control males. Furthermore, there was a significant difference in progeny survival between the probiotic-fed group and the control group at three developmental times (24 hours postfertilization (hpf), 5 days postfertilization (dpf) and 7 dpf). In conclusion, in zebrafish, prolonged ingestion of a mixture of Lactobacillus rhamnosus CECT8361 and Bifidobacterium longum CECT7347 has positive effects on growth, sperm quality, and progeny survival.

Diet Supplemented with Antioxidant and Anti-Inflammatory Probiotics Improves Sperm Quality after Only One Spermatogenic Cycle in Zebrafish Model.

Infertility is a medical concern worldwide and could also have economic consequences in farmed animals. Developing an efficient diet supplement with immediate effects on sperm quality is a promising tool for human reproduction and for domesticated animal species. This study aims at elucidating the effect of a short-time probiotic supplementation consisting of a mixture of two probiotic bacteria with proven antioxidant and anti-inflammatory activities on zebrafish sperm quality and male behavior. For this purpose, three homogeneous groups of males in terms of motility (<60%) were established. The control group was fed with a normal standard diet. The other received supplements: One group (vehicle control) was fed with maltodextrin and the other received a probiotic preparation based on a mixture (1:1) of Lactobacillus rhamnosus CECT8361 and Bifidobacterium longum CECT7347. The feeding regime was 21 days corresponding with a single spermatogenesis in zebrafish. The preparation did not modify animal weight, positively affected the number of fluent males, increased sperm concentration, total motility, progressive motility, and fast spermatozoa subpopulations. Moreover, the animals fed with the supplement showed different behavior patterns compared to control groups. Our results suggest a diet-related modulation on the exploration activity indicating a lower stress-like conduct. The studied formulation described here should be considered as advantageous in male reproductive biotechnology.

Artificial Neural Network (ANN) as a Tool to Reduce Human-Animal Interaction Improves Senegalese Sole Production.

Manipulation is usually required for biomass calculation and food estimation for optimal fish growth in production facilities. However, the advances in computer-based systems have opened a new range of applied possibilities. In this study we used image analysis and a neural network algorithm that allowed us to successfully provide highly accurate biomass data. This developed system allowed us to compare the effects of reduced levels of human-animal interaction on the culture of adult Senegalese sole (Soleasenegalensis) in terms of body weight gain. For this purpose, 30 adult fish were split into two homogeneous groups formed by three replicates (n=5) each: a control group (CTRL), which was standard manipulated and an experimental group (EXP), which was maintained under a lower human-animal interaction culture using our system for biomass calculation. Visible implant elastomer was, for the first time, applied as tagging technology for tracking soles during the experiment (four months). The experimental group achieved a statistically significant weight gain (p<0.0100) while CTRL animals did not report a statistical before-after weight increase. Individual body weight increment was lower (p<0.0100) in standard-handled animals. In conclusion, our experimental approach provides evidence that our developed system for biomass calculation, which implies lower human-animal interaction, improves biomass gain in Senegalese sole individuals in a short period of time.