Sperm functionality is differentially regulated by porcine oviductal extracellular vesicles from the distinct phases of the estrous cycle.

Context Extracellular vesicles (EVs) derived from the oviductal fluid (oEVs) play a critical role in various reproductive processes, including sperm capacitation, fertilisation, and early embryo development. Aims To characterise porcine oEVs (poEVs) from different stages of the estrous cycle (late follicular, LF; early luteal, EL; mid luteal, ML; late luteal, LL) and investigate their impact on sperm functionality. Methods poEVs were isolated, characterised, and labelled to assess their binding to boar spermatozoa. The effects of poEVs on sperm motility, viability, acrosomal status, protein kinase A phosphorylation (pPKAs), tyrosine phosphorylation (Tyr-P), and in in vitro fertility were analysed. Key results poEVs were observed as round or cup-shaped membrane-surrounded vesicles. Statistical analysis showed that poEVs did not significantly differ in size, quantity, or protein concentration among phases of the estrous cycle. However, LF poEVs demonstrated a higher affinity for binding to sperm. Treatment with EL, ML, and LL poEVs resulted in a decrease in sperm progressive motility and total motility. Moreover, pPKA levels were reduced in presence of LF, EL, and ML poEVs, while Tyr-P levels did not differ between groups. LF poEVs also reduced sperm penetration rate and the number of spermatozoa per penetrated oocyte (P Conclusions poEVs from different stages of the estrous cycle play a modulatory role in sperm functionality by interacting with spermatozoa, affecting motility and capacitation, and participating in sperm-oocyte interaction. Implications The differential effects of LF and LL poEVs suggest the potential use of poEVs as additives in IVF systems to regulate sperm-oocyte interaction.

Artificial insemination of all ejaculated sperm fractions accelerates embryo development and increases the uterine vascularity in the pig.

The semen of boar is characterized by ejaculation in well-differentiated fractions with specific concentration, composition, and volume. The 'sperm-rich fraction' (SRF), the most concentrated seminal fraction, is habitually collected in insemination centers to make artificial insemination (AI) doses. The absence of the other fractions in AI doses could alter the uterine reaction to AI and not trigger essential responses that could maximize fertility. Thus, there is an urge to ascertain the impact of different ejaculate fractions on the uterus after AI to optimize the semen doses. This work analyzed specific parameters related to fertility in pregnant artificially inseminated sows (n = 15) with ac-cumulative fractions of the semen of boars (n = 6): F1, composed of the sperm-rich fraction (SRF); F2, composed of F1 plus the intermediate fraction; F3, composed of F2 plus the post-SRF. Non-inseminated sows (n = 5) were included as control (C). The different types of seminal dose did not affect the number of ovulated follicles (CL; corpora lutea, p > 0.05) but did affect the embryo development (p < 0.05). The proportion of embryos in morula stages was significantly higher in AI-F1 sows (84.4%, p < 0.05). Morulas and blastocysts were balanced in AI-F2 or AI-F3 (p > 0.05). Independently of the type of seminal dose (F1, F2, or F3), we observed by immunohistochemistry that AI significantly increased uterine vascularization, although with some anatomical differences. The cranial region of the uterine horns was significantly more vascularized in AI-F1 or AI-F2 sows (26.7 ± 2.3 and 28.6 ± 2.0%, respectively), and AI-F3 showed significantly less vascularization at that point (17.8 ± 1.6%, p < 0.05). To summarize, the synergistic effect of all ejaculate fractions accelerates embryo development, at least during the preimplantation period, and increases the uterine reaction to AI in certain parts of the uterus.

Different seminal ejaculated fractions in artificial insemination condition the protein cargo of oviductal and uterine extracellular vesicles in pig.

The seminal plasma (SP) is the liquid component of semen that facilitates sperm transport through the female genital tract. SP modulates the activity of the ovary, oviductal environment and uterine function during the periovulatory and early pregnancy period. Extracellular vesicles (EVs) secreted in the oviduct (oEVs) and uterus (uEVs) have been shown to influence the expression of endometrial genes that regulate fertilization and early embryo development. In some species, semen is composed of well-separated fractions that vary in concentration of spermatozoa and SP composition and volume. This study aimed to investigate the impact of different accumulative fractions of the porcine ejaculate (F1, composed of the sperm-rich fraction, SRF; F2, composed of F1 plus the intermediate fraction; F3, composed of F2 plus the post-SRF) on oEVs and uEVs protein cargo. Six days after the onset of estrus, we determined the oEVs and uEVs size and protein concentration in pregnant sows by artificial insemination (AI-sows) and in non-inseminated sows as control (C-sows). We also identified the main proteins in oEVs and uEVs, in AI-F1, AI-F2, AI-F3, and C-sows. Our results indicated that although the size of EVs is similar between AI- and C-sows, the protein concentration of both oEVs and uEVs was significantly lower in AI-sows (p < 0.05). Proteomic analysis identified 38 unique proteins in oEVs from AI-sows, mainly involved in protein stabilization, glycolytic and carbohydrate processes. The uEVs from AI-sows showed the presence of 43 unique proteins, including already-known fertility-related proteins (EZR, HSPAA901, PDS). We also demonstrated that the protein composition of oEVs and uEVs differed depending on the seminal fraction(s) inseminated (F1, F2, or F3). In conclusion, we found specific protein cargo in oEVs and uEVs according to the type of semen fraction the sow was inseminated with and whose functions these specific EVs proteins are closely associated with reproductive processes.

Low-density colloid centrifugation removes bacteria from boar semen doses after spiking with selected species.

Single-layer centrifugation (SLC) with a low-density colloid is an efficient method for removing contaminating microorganisms from boar semen while recovering most spermatozoa from the original sample. This study tested the performance of this technique, using 50-ml tubes, by spiking commercial semen doses prepared without antibiotics with selected bacterial species followed by storage at 17 °C. The doses were spiked up to 102/ml CFU (colony forming units) of the bacteria Burkholderia ambifaria, Pseudomonas aeruginosa, and Staphylococcus simulans. The semen was processed by SLC (15 ml of sample and 15 ml of colloid) with the colloid Porcicoll at 20% (P20) and 30% (P30), with a spiked control (CTL) and an unspiked control (CTL0), analyzing microbiology and sperm quality on days 0, 3 and 7. SLC completely removed B. ambifaria and S. simulans, considerably reducing P. aeruginosa and overall contamination (especially P30, ∼104 CFU/ml of total contamination on day 7, median). Sperm viability was lower in P20 and P30 samples at day 0, with higher cytoplasmic ROS. Still, results were similar in all groups on day 3 and reversed on day 7, indicating a protective effect of SLC (possibly directly by removal of damaged sperm and indirectly because of lower bacterial contamination). Sperm chromatin was affected by the treatment (lower DNA fragmentation and chromatin decondensation) and storage (higher overall condensation on day 7 as per chromomycin A3 and monobromobimane staining). In conclusion, SLC with low-density colloids can remove most bacteria in a controlled contamination design while potentially improving sperm quality and long-term storage at practical temperatures.

Setting up regional diagnostic reference levels for pediatric interventional cardiology in Latin America and the Caribbean countries: preliminary results and identified challenges.

The goal of the present study was to propose a set of preliminary regional diagnostic reference levels (DRLs) for pediatric interventional cardiology (IC) procedures in Latin America and the Caribbean countries, classified by age and weight groups. The study was conducted in the framework of the Optimization of Protection in Pediatric Interventional Radiology in Latin America and the Caribbean program coordinated by the World Health Organization and the Pan American Health Organization in cooperation with the International Atomic Energy Agency. The first step of the program was focused on pediatric IC. Dose data from diagnostic and therapeutic procedures were collected between December 2020 and December 2021. Regional DRLs were set as the third quartile of patient dose data (kerma area product) collected in 18 hospitals from 10 countries in an initial sample of 968 procedures. DRLs were set for four age bands and five weight ranges. The values obtained for the four age bands (<1 yr, 1 to <5 yr, 5 to <10 yr and 10 to <16 yr) were 2.9, 6.1, 8.8 and 14.4 Gy cm2for diagnostic procedures, and 4.0, 5.0, 10.0 and 38.1 Gy cm2for therapeutic procedures, respectively. The values obtained for the five weight bands (<5 kg, 5 to <15 kg, 15 to <30 kg, 30 to <50 kg and 50 to <80 kg) were 3.0, 4.5, 8.1, 9.2 and 26.8 Gy cm2for diagnostic procedures and 3.7, 4,3, 7.3, 16.1 and 53.4 Gy cm2for therapeutic procedures, respectively. While initial data were collected manually as patient dose management systems (DMSs) were not available in most of the hospitals involved in the program, a centralized automatic DMS for the collection and management of patient dose indicators has now been introduced and is envisaged to increase the sample size. The possibility of alerting on high dose values and introducing corrective actions will help in optimization.

Reproductive fluids, used for the in vitro production of pig embryos, result in healthy offspring and avoid aberrant placental expression of PEG3 and LUM.

BACKGROUND: In vitro embryo production (IVP) and embryo transfer (ET) are two very common assisted reproductive technologies (ART) in human and cattle. However, in pig, the combination of either procedures, or even their use separately, is still considered suboptimal due to the low efficiency of IVP plus the difficulty of performing ET in the long and contorted uterus of the sow. In addition, the potential impact of these two ART on the health of the offspring is unknown. We investigated here if the use of a modified IVP system, with natural reproductive fluids (RF) as supplements to the culture media, combined with a minimally invasive surgery to perform ET, affects the output of the own IVP system as well as the reproductive performance of the mother and placental molecular traits. RESULTS: The blastocyst rates obtained by both in vitro systems, conventional (C-IVP) and modified (RF-IVP), were similar. Pregnancy and farrowing rates were also similar. However, when compared to in vivo control (artificial insemination, AI), litter sizes of both IVP groups were lower, while placental efficiency was higher in AI than in RF-IVP. Gene expression studies revealed aberrant expression levels for PEG3 and LUM in placental tissue for C-IVP group when compared to AI, but not for RF-IVP group. CONCLUSIONS: The use of reproductive fluids as additives for the culture media in pig IVP does not improve reproductive performance of recipient mothers but could mitigate the impact of artificial procedures in the offspring.

Volatile metabolites produced by different flor yeast strains during wine biological ageing.

Sherry white wine called Fino is produced by dynamic biological ageing under the action of flor yeasts using traditional practices aimed at ensuring uniform quality and characteristics over time. These kinds of yeasts provide typical sensory properties to Fino wines. Although there are studies of the volatile composition of these wines submitted to biological ageing in wood barrels, there is a lack of knowledge on the particular volatile profile produced by different flor yeast strains from Sherry zone wineries. For this reason, the aim of this study was to analyse the volatile profiles produced by 15 pure culture flor velum yeasts, with the goal of observing their suitability for obtaining high quality Fino sherry wines. Volatile composition was determined by dual sequential stir bar sorptive extraction, followed by GC-MS analysis. All yeast strains studied produced the increase of most acetals, highlighting acetaldehyde diethylacetal which was the compound that most increased. Among terpenes, nerolidol and farnesol underwent remarkable increases. However, results showed that in a month of biological ageing, significant differences were observed among the volatile metabolites produced by flor yeast strains studied. Only some of them stood out for their high production of volatile compounds characteristic of Sherry Fino wines, which are good candidates for producing starter cultures.

Eye lens radiation exposure in paediatric interventional treatment of retinoblastoma.

Retinoblastoma represents 3% of cancers in children under fifteen years of age. The standard paediatric treatment for saving the affected eye is supraselective intra-arterial chemotherapy performed in interventional rooms. In order to address the radiation toxicity due to the angiography, the aim of this study was to determine the typical dose value corresponding to the procedure, estimate the paediatric patients' eye lens dose and study the relationship between dose indicators and dose to the lens. An automatic dose management software was installed in two interventional rooms to obtain the distribution of the dose indicators kerma-area product and reference-point air kerma, getting a typical value 16 Gy·cm2 and 130 mGy, respectively (n = 35). The eye lens dose estimates were obtained with photoluminescent dosimeters placed on the patient's eyelids. In the left eye, the entrance surface air kerma was 44.23 ± 2.66 mGy, and 12.72 ± 0.89 mGy in the right eye (n = 10). There was a positive correlation between dose to the lens per procedure and dose indicators, with R2 > 0.65 for both eyes. Based on this information, the threshold for the onset of radiation-induce cataracts (500 mGy) will be exceeded if the treatment is performed for more than 8 sessions.

Identification of volatile biomarkers of Giardia duodenalis infection in children with persistent diarrhoea.

Currently, chronic diarrhoea syndrome in children is a very common pathology whose aetiology is sometimes difficult to identify. Methodologies for the diagnosis of infections have diversified, and gas chromatography/mass spectrometry (GC/MS) is a very useful tool. The aim of this study was to identify volatile biomarkers of the presence of Giardia duodenalis in the faeces of patients with chronic diarrhoea (with and without giardiasis) using static headspace extraction followed by GC/MS. The analysis of the volatiles extracted from the headspace had enough sensitivity to detect differences in the volatile profiles in the faeces of the patients with and without Giardia duodenalis infection and discriminate between them. Dimethyl disulphide and trisulphide were found in the faeces of patients without giardiasis but not in the faeces of patients with G. duodenalis. Finally, three possible biomarkers, acetic acid, 1,4-dimethoxy-2,3-butanediol and 1,3-dimethoxy-2-propanol, were proposed to identify patients with giardiasis; these compounds were not present in the patients without the parasite. Multivariate analysis revealed that principal component 1 separated the stool samples according to the presence of infection by G. duodenalis despite the inter-individual variability in biological specimens such as faeces.

Tissue plasminogen activator (tPA) of paternal origin is necessary for the success of in vitro but not of in vivo fertilisation in the mouse.

Besides its fibrinolytic function, the plasminogen-plasmin (PLG-PLA) system is also involved in fertilisation, where plasminogen activators bind to plasminogen to produce plasmin, which modulates sperm binding to the zona pellucida. However, controversy exists, depending on the species, concerning the role of the different components of the system. This study focused its attention on the role of the PLG-PLA system on fertilisation in the mouse with special attention to tissue plasminogen activator (tPA). The presence of exogenous plasminogen reduced invitro fertilisation (IVF) rates and this decline was attenuated by the presence of plasmin inhibitors in combination with plasminogen. The incubation of spermatozoa with either oocytes or cumulus cells together with plasminogen did not change the acrosome reaction but reduced the number of spermatozoa attached. When spermatozoa from tPA-/- mice were used, the IVF rate decreased drastically, although the addition of exogenous tPA during gamete co-incubation under invitro conditions increased fertilisation success. Moreover, fertility could not be restored after invivo insemination of tPA-/- spermatozoa in the female ampulla, although tPA-/- males were able to fertilise invivo. This study suggests a regulatory role of the PLG-PLA system during fertilisation in the mouse with possible implications in human reproduction clinics, such as failures in tPA production, which could be partially resolved by the addition of exogenous tPA during IVF treatment.

Influence of Saccharomyces cerevisiae and Lachancea thermotolerans co-inoculation on volatile profile in fermentations of a must with a high sugar content.

The aim of this work was to evaluate how the use of mixed cultures of Saccharomyces cerevisiae and Lachancea thermotolerans indigenous yeast strains influences the volatile composition of wine. Multivariate curve resolution (MCR) method has been applied to data analysis. Five fermentation trials were carried out: three co-inoculated with L. thermotolerans:S. cerevisiae, at the ratio of 50:1, 20:1 and 5:1 respectively, and two with a pure culture of each strain. A must from sun-dried Pedro Ximénez grapes was employed. Volatile composition was determined by dual sequential stir bar sorptive extraction, followed by GC-MS analysis. MCR provided 171 peaks. Results in co-inoculation fermentations revealed that the majority compounds obtained in each case followed the same trend as that observed for the pure culture of the strain that was present in a higher proportion. Finally, 50:1 and 20:1 seemed to be the best proportions to obtain a wine with a quality volatile profile.

Application of newly developed Fluoro-QC software for image quality evaluation in cardiac X-ray systems.

INTRODUCTION: A quality assurance (QA) program is a valuable tool for the continuous production of optimal quality images. The aim of this paper is to assess a newly developed automatic computer software for image quality (IR) evaluation in fluoroscopy X-ray systems. METHODS: Test object images were acquired using one fluoroscopy system, Siemens Axiom Artis model (Siemens AG, Medical Solutions Erlangen, Germany). The software was developed as an ImageJ plugin. Two image quality parameters were assessed: high-contrast spatial resolution (HCSR) and signal-to-noise ratio (SNR). The time between manual and automatic image quality assessment procedures were compared. The paired t-test was used to assess the data. p Values of less than 0.05 were considered significant. RESULTS: The Fluoro-QC software generated faster IQ evaluation results (mean = 0.31 ± 0.08 min) than manual procedure (mean = 4.68 ± 0.09 min). The mean difference between techniques was 4.36 min. Discrepancies were identified in the region of interest (ROI) areas drawn manually with evidence of user dependence. The new software presented the results of two tests (HCSR = 3.06, SNR = 5.17) and also collected information from the DICOM header. Significant differences were not identified between manual and automatic measures of SNR (p value = 0.22) and HCRS (p value = 0.46). CONCLUSION: The Fluoro-QC software is a feasible, fast and free to use method for evaluating imaging quality parameters on fluoroscopy systems.

Paediatric interventional cardiology in Costa Rica: diagnostic reference levels and estimation of population dose.

The goal of the present study was to propose a set of national diagnostic reference levels (DRLs) in Costa Rica for paediatric interventional cardiology (IC) procedures classified by age and weight and to estimate the collective dose of the paediatric population from these intervention practices. The data collection period was May 2016 to May 2017. The third quartile of patient dose data distributions for kerma-area product (KAP) values was set as the national DRLs. A sample of 154 paediatric IC procedures (collected in the national paediatric hospital with a single x-ray system) was used and divided into four age ranges and five weight ranges. The national DRLs obtained for KAPs by age range were 1.79 Gy cm2 (<1 year) to 23.0 Gy cm2 (10-15 years). The national DRLs obtained for KAPs by weight range were 1.0 Gy cm2 (<10 kg) to 49.6 Gy cm2 (50-79 kg). The contribution to the collective dose of the population of Costa Rica amounted to 0.78 person Sv.

Differential gene expression in porcine oviduct during the oestrous cycle.

The oviduct undergoes changes under the influence of steroid hormones during the oestrous cycle. However, the molecular mechanisms underlying oviductal regulation are not fully understood. The aim of the present study was to identify the gene expression profile of the porcine oviduct in different stages of the cycle using microarray technology. A systematic study was performed on animals at four different stage: prepubertal gilts, and sows in the preovulatory, postovulatory and luteal phase of the oestrous cycle. The porcine oviduct expressed a total of 4929 genes. Moreover, significant differences in the expression of several genes were detected as the oestrous cycle progressed. Analysis of the differentially expressed genes indicated that a total of 86, 89 and 15 genes were upregulated in prepubertal gilts, preovulatory and luteal sows respectively compared with levels observed in postovulatory sows. Moreover, 80, 51 and 64 genes were downregulated in prepubertal, preovulatory and luteal animals respectively compared with the postovulatory sows. The concentrations of 10 selected transcripts were quantified by real-time reverse transcription-polymerase chain reaction to validate the cDNA array hybridisation data. Conversely, for some genes, localisation of corresponding protein expression in the oviduct was analysed by immunohistochemistry (i.e. cholecystokinin, glutathione peroxidase 2, mucin 1, phosphatidylethanolamine binding protein 4 and tachykinin 3) and mass spectrometry analysis of oviductal fluid allowed identification of peptides from all five proteins. The results of the present study demonstrate that gene expression in the porcine oviduct is clearly regulated during the oestrous cycle, with some oviductal proteins that could be related to several reproductive processes described here for the first time.

The C-terminal region of OVGP1 remodels the zona pellucida and modifies fertility parameters.

OVGP1 is the major non-serum glycoprotein in the oviduct fluid at the time of fertilization and early embryo development. Its activity differs among species. Here, we show that the C-terminal region of recombinant OVGP1 regulates its binding to the extracellular zona pellucida and affects its activity during fertilization. While porcine OVGP1 penetrates two-thirds of the thickness of the zona pellucida, shorter OVGP1 glycoproteins, including rabbit OVGP1, are restricted to the outer one-third of the zona matrix. Deletion of the C-terminal region reduces the ability of the glycoprotein to penetrate through the zona pellucida and prevents OVGP1 endocytosis. This affects the structure of the zona matrix and increases its resistance to protease digestion. However, only full-length porcine OVGP1 is able to increase the efficiency rate of in vitro fertilization. Thus, our findings document that the presence or absence of conserved regions in the C-terminus of OVGP1 modify its association with the zona pellucida that affects matrix structure and renders the zona matrix permissive to sperm penetration and OVGP1 endocytosis into the egg.

Volatile profile characterisation of Chilean sparkling wines produced by traditional and Charmat methods via sequential stir bar sorptive extraction.

The volatile compositions of Charmat and traditional Chilean sparkling wines were studied for the first time. For this purpose, EG-Silicone and PDMS polymeric phases were compared and, afterwards, the most adequate was selected. The best extraction method turned out to be a sequential extraction in the headspace and by immersion using two PDMS twisters. A total of 130 compounds were determined. In traditional Chilean sparkling wines, ethyl esters were significantly higher, while acetic esters and ketones were predominant in the Charmat wines. PCA and LDA confirmed the differences in the volatile profiles between the production methods (traditional vs. Charmat).

Meningitis por Listeria monocytogenes en paciente inmunocompetente / Listeria monocytogenes meningitis in immunocompetent patient

Listeria monocytogenes es un patógeno de origen alimentario que suele producir gastroenteritis, procesos febriles, sepsis y meningitis. Afecta característicamente a neonatos, embarazadas, ancianos e inmunocomprometidos, con una epidemiología controvertida y poco conocida. Se presenta un caso de meningitis y síndrome de secreción inadecuada de hormona antidiurética secundario en paciente inmunocompetente (AU)

Listeria monocytogenes is a foodborne pathogen that usually cause gastroenteritis, fever, sepsis and meningitis, which characteristically affects immunocompromised, newborns, pregnant women and elderly people, with controversial and unknown epidemiology. We report a case of meningitis and secondary inadequate secretion of antidiuretic hormone syndrome in immunocompetent patient (AU)

Recent developments in the analysis of musty odour compounds in water and wine: A review.

One of the most common taints in foods is a musty or earthy odour, which is commonly associated with the activity of microorganisms. Liquid foods, particularly wine and water, can be affected by this defect due to the presence of certain aromatic organic compounds at very low concentrations (ng/L) consistent with human threshold perception levels. The volatile compounds responsible for a mouldy off-aroma include approximately 20 compounds, namely, haloanisoles, geosmin, 2-methylisoborneol, several alkyl-methoxypyrazines, 1-octen-3-ol, 1-octen-3-one, trans-octenol, 3-octanone, fenchol and fenchone. Methods for determining these very low concentrations of odour compounds must be extremely sensitive and selective with efficient preconcentration treatments. A number of extraction techniques based on LLME (liquid-liquid microextraction), SPME (solid-phase microextraction) or SBSE (stir-bar sorptive extraction) can be applied and should be selected on a case-by-case basis. Moreover, recent developments in GC instrumentation coupled to different detection systems can effectively increase the selectivity and sensitivity of the analysis of target compounds.