Proteomic profile of vitrified in vitro-produced bovine embryos (Bos taurus indicus).

BACKGROUND: The proteomic profile of cryopreserved in vitro produced bovine embryos is little known but can provide insights on the successful application of cryo procedures in support of animal breeding. OBJECTIVE: To identify embryonic proteins and biomarkers related to improved cryotolerance of vitrified in vitro produced bovine embryos. MATERIALS AND METHODS: Proteins were isolated from embryo pools (n = 25 embryos per replicate) and analyzed using the nanoLC - MS/MS system. Further, the UniProtKB database (Uniprot -http://www.uniprot.org/) was used for protein identification. Proteins were classified based on their molecular mass, isoelectric point, and enzymatic activity. Post-translational modification predictions and functional gene ontology analysis were performed as well. Finally, a protein-protein interaction network was created to shed light on the embryo interactome. RESULTS: Based on the MS/MS approach, 66 proteins were identified from vitrified Bos taurus embryos. The retrieved proteins were presumably annotated, which allowed a description of the qualitative and functional aspects of the embryo proteome after the vitrification process. CONCLUSION: These findings allowed us to conclude that in vitro-produced vitrified embryos expressed proteins that underlie biological processes related to reproduction, stress and lipid metabolic process, which are essential to maintain embryo viability. doi.org/10.54680/fr22410110512.

Characterizing network topology using first-passage analysis.

Understanding the topological characteristics of complex networks and how they affect navigability is one of the most important goals in science today, as it plays a central role in various economic, biological, ecological, and social systems. Here we apply first-passage analysis tools to investigate the properties and characteristics of random walkers in networks with different topology. Starting with the simplest two-dimensional square lattice, we modify its topology incrementally by randomly reconnecting links between sites. We characterize these networks by first-passage time from a significant number of random walkers without interaction, varying the departure and arrival locations. We also apply the concept of first-passage simultaneity, which measures the likelihood of two walkers reaching their destination together. These measures, together with the site occupancy statistics during the processes, allowed us to differentiate the studied networks, especially the random networks from the scale-free networks, by their navigability. We also show that small-world features can also be highlighted with the proposed technique.

Evaluation of morphology, morphometry and follicular dynamics in FecGE genotyped ewes.

This study aimed to evaluate the effect of FecGE mutation on the development of ovarian follicles. To this end, 42 Santa Inês ewes were genotyped for FecGE mutation and classified as wild-type (FecG+/+), heterozygous (FecG+/E) or mutant homozygous (FecGE/E). Ovarian fragments were processed, and the follicles were analyzed with regard to the morphology and morphometry using classical histology. For the evaluation of follicular dynamics, ewes underwent oestrous synchronization and were monitored throughout an interovulatory period. A higher (P < 0.05) percentage of morphologically normal follicles in the primordial stage was identified in FecGE/E (90.0%) and FecG+/E (88.1%) ewes than in the FecG+/+ (73.0%) ewes. There was also a significantly greater (P < 0.05) number of morphologically normal follicles in the FecGE/E (87.3%) and FecG+/E (83.3%) ewes than in FecG+/+ (76.8%) ewes in the transitional stage. A smaller (P < 0.05) diameter was observed in the secondary follicles in FecGE/E (93.8 µm) ewes than in FecG+/E (171.8 µm) ewes. Regarding follicular dynamics, FecGE/E ewes showed a greater (P < 0.05) number of ovulations (2.5 ±â€¯0.2) than FecG+/+ ewes (1.5 ±â€¯0.3) ewes. Ovulatory follicles were smaller (P < 0.05) in the FecGE/E (5.1 mm) and FecG+/E (5.2 mm) ewes than in FecG+/+ (5.8 mm) ewes. Santa Inês nulliparous ewes carrying the FecGE mutation showed a greater proportion of morphologically normal follicles in the primordial and transitional stages than those not carrying the mutation. FecGE/E ewes demonstrated a higher number of ovulated follicles and that FecGE/E and FecG+/E ewes presented ovulatory follicles with a smaller diameter.

In Vitro and In Vivo Efficiency of Extenders Added to Thawed Stallion Semen.

BACKGROUND: Addition of extenders to thawed semen could improve fertility. OBJECTIVE: To determine the efficiency of extenders to increase viability of thawed semen, measured by sperm parameters in vitro and pregnancy rates after artificial insemination (AI). MATERIALS AND METHODS: Sperm motility and acrosin activity were measured during a thermoresistance test (TRT). RESULTS: Progressive motility decreased (P<0.05) after 30 min in thawing semen treated with saline solution (SS) and only after 60 min with Tyrode's solution (TS) or freezing diluent (FD). The total motility decreased (P<0.05) after 60 min in thawed semen treated with SS, and after 90 min in thawed semen containing TS or FD. The acrosin activity decreased (P<0.05) after 60 min during the TRT, but there was no difference among treatments throughout the TRT. The pregnancy rates were similar among thawed-semen supplemented with SS, TS or FD. CONCLUSION: The extenders neither improve sperm parameters nor enhance AI results.

Mucormycosis outbreak due to Rhizopus microsporus after arthroscopic anterior cruciate ligament reconstruction surgery evaluated by RAPD and MALDI-TOF Mass spectrometry.

BACKGROUND: Rhizopus microsporus is one of the main causative agents of mucormycosis. These mycoses are mostly described as isolated cases involving uncontrolled diabetes mellitus or immunosuppressed patients. In this work we report a nosocomial outbreak of mucormycosis due to R. microsporum involving three young immunocompetent patients whom underwent arthroscopic anterior cruciate ligament reconstruction surgery in a seven-month time span. PROCEDURES: During the outbreak period, a total of 32 surgeries of this type were performed in the clinic (mucormycosis prevalence of 9.375%). The three patients presented healthcare-associated Mucormycosis comprising the bone surrounding one of the fixation screws (femoral or tibial). In addition to these three strains, another three R. microsporus strains isolated in the medical center during the same period of time were included in the study. One of these fungi was isolated from a skin lesion of a kidney transplant patient while the other two strains were isolated from environmental sources. Classical, mass spectrometry-based (MALDI-TOFF) and molecular identification were performed. Genetic relatedness was established by Rep-PCR (RAPD variant) and by single-linkage cluster analysis mass spectra. Cluster analysis was performed by unweighed pair group method with arithmetic mean (UPGMA). MAIN FINDINGS: All the strains were identified as R. microsporum by the used phenotypic and genetic tools. Clinical strains fell into 2 different clusters separating the renal transplant recipient strain from the three strains isolated post ACLR surgery, which clustered together. CONCLUSIONS: The established genetic/mass spectra relatedness between the three post-surgery isolates suggests that these cases may be considered a healthcare-associated mucormycosis outbreak.

Temporal expression of pluripotency-associated transcription factors in sheep and cattle preimplantation embryos.

SummaryPluripotency-associated transcription factors (PATFs) modulate gene expression during early mammalian embryogenesis. Despite a strong understanding of PATFs during mouse embryogenesis, limited progress has been made in ruminants. This work aimed to describe the temporal expression of eight PATFs during both sheep and cattle preimplantation development. Transcript availability of PATFs was evaluated by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) in eggs, cleavage-stage embryos, morulae, and blastocysts. Transcripts of five genes were detected in all developmental stages of both species (KLF5, OCT4, RONIN, ZFP281, and ZFX). Furthermore, CMYC was detected in all cattle samples but was found from cleavage-stage onwards in sheep. In contrast, NR0B1 was detected in all sheep samples but was not detected in cattle morulae. GLIS1 displayed the most significant variation in temporal expression between species, as this PATF was only detected in cattle eggs and sheep cleavage-stage embryos and blastocysts. In silico analysis suggested that cattle and sheep PATFs share similar size, isometric point and molecular weight. A phenetic analysis showed two patterns of PATF clustering between cattle and sheep, among several mammalian species. In conclusion, the temporal expression of pluripotency-associated transcription factors differs between sheep and cattle, suggesting species-specific regulation during preimplantation development.

Site of intrauterine artificial insemination in the bitch does not affect sperm distribution within the uterus.

The aim of this study was to evaluate the distribution of frozen-thawed spermatozoa within the uterine lumen and oviducts following intrauterine laparoscopic deposition at two sites. Twelve bitches of unknown reproductive history were randomly distributed into two groups. Semen (3 ml containing 300 × 10(6) frozen-thawed spermatozoa) was infused at the uterine body (UB group) or at the cranial tip of the left uterine horn. A 22-G catheter was used to access the uterine lumen. Sperm cell distribution was evaluated after ovariohysterectomy performed 3 h after artificial insemination (AI). There was no difference between groups in mean time to perform AI. Spermatozoa were detected in all uterine segments, including the tip of both horns, but none was detected in the oviduct. The 22-G catheter facilitated deposition of semen in the uterine lumen, particularly at the UB site. Sperm cell distribution occurred evenly along both horns, independent of the site of semen deposition.

Virulence Variability of Puccinia coronata f. sp. avenae Isolates Collected in Three Counties from Rio Grande do Sul State, Brazil.

Using isolates collected in three counties of Rio Grande do Sul State, Brazil, the goals of this work were to determine (i) the pattern of virulence or avirulence of the isolates to 25 Pc resistance genes, (ii) the similarity in virulence among Puccinia coronata f. sp. avenae isolates considering their pattern of virulence or avirulence, (iii) the race code for each isolate by the North American system of nomenclature, and (iv) the supplemental Pc genes potentially useful as local differentials for P. coronata f. sp. avenae races. The results indicate that the southern Brazilian rust isolates presented a high level of virulence, because 66% of inoculations manifested the high infection type. Only the Pc 68 gene was effective against all tested isolates. In general, each isolate presented a different pattern of virulence or avirulence, which indicates the high variability for virulence that the fungus presents at the sampled sites. However, the North American system of nomenclature was not completely sufficient in distinguishing southern Brazilian races. Thus, the genes Pc 36, Pc 53, Pc 55, and Pc 63 represent a possible gene combination to be incorporated into the North American system of nomenclature.